bims-bicyki 2022-07-31 papers

bims-bicyki

Biomed News

on Bicaudal-C1 and interactors in cystic kidney disease

Issue of 2022‒07‒31
twenty-one papers selected by
Céline Gagnieux
École Polytechnique Fédérale de Lausanne (EPFL)

1-Indanone retards cyst development in ADPKD mouse model by stabilizing tubulin and down-regulating anterograde transport of cilia.
The Pathophysiology of Left Ventricular Hypertrophy, beyond Hypertension, in Autosomal Dominant Polycystic Kidney Disease.
Modulation of Primary Cilia by Alvocidib Inhibition of CILK1.
Non-cell-autonomous activation of Hedgehog signaling contributes to disease progression in a mouse model of renal cystic ciliopathy.
Monoallelic pathogenic ALG5 variants cause atypical polycystic kidney disease and interstitial fibrosis.
A 3D In Vivo Model for Studying Human Renal Cystic Tissue and Mouse Kidney Slices.
Polycystin-2 in the Endoplasmic Reticulum: Bending Ideas about the Role of the Cilium.
Disruption of dopamine receptor 1 localization to primary cilia impairs signaling in striatal neurons.
Manipulation of the Tubulin Code Alters Directional Cell Migration and Ciliogenesis.
TTC30A and TTC30B Redundancy Protects IFT Complex B Integrity and Its Pivotal Role in Ciliogenesis.
Deep Learning Automation of Kidney, Liver, and Spleen Segmentation for Organ Volume Measurements in Autosomal Dominant Polycystic Kidney Disease.
Trophectoderm formation: regulation of morphogenesis and gene expressions by RHO, ROCK, cell polarity, and HIPPO signaling.
Arih2 regulates hedgehog signaling through smoothened ubiquitination and ER-associated degradation.
Ciliogenesis requires sphingolipid-dependent membrane and axoneme interaction.
Evidence of nonsurgical treatment for polycystic liver disease.
Regulated processing and secretion of a peptide precursor in cilia.
Strongly Truncated Dnaaf4 Plays a Conserved Role in Drosophila Ciliary Dynein Assembly as Part of an R2TP-Like Co-Chaperone Complex With Dnaaf6.
miR449 Protects Airway Regeneration by Controlling AURKA/HDAC6-Mediated Ciliary Disassembly.
Retinal Ciliopathy in the Patient with Transplanted Kidney: Case Report.
Identical IFT140 Variants Cause Variable Skeletal Ciliopathy Phenotypes-Challenges for the Accurate Diagnosis.
Challenges in Diagnosing Primary Ciliary Dyskinesia in a Brazilian Tertiary Hospital.

Acta Pharmacol Sin. 2022 Jul 29.

1-Indanone retards cyst development in ADPKD mouse model by stabilizing tubulin and down-regulating anterograde transport of cilia.

Xiao-Wei Li, Jian-Hua Ran, Hong Zhou, Jin-Zhao He, Zhi-Wei Qiu, Shu-Yuan Wang, Meng-Na Wu, Shuai Zhu, Yong-Pan An, Ang Ma, Min Li, Ya-Zhu Quan, Nan-Nan Li, Chao-Qun Ren, Bao-Xue Yang.

  Autosomal dominant polycystic kidney disease (ADPKD) is the most common inherited kidney disease. Cyst development in ADPKD involves abnormal epithelial cell proliferation, which is affected by the primary cilia-mediated signal transduction in the epithelial cells. Thus, primary cilium has been considered as a therapeutic target for ADPKD. Since ADPKD exhibits many pathological features similar to solid tumors, we investigated whether targeting primary cilia using anti-tumor agents could alleviate the development of ADPKD. Twenty-four natural compounds with anti-tumor activity were screened in MDCK cyst model, and 1-Indanone displayed notable inhibition on renal cyst growth without cytotoxicity. This compound also inhibited cyst development in embryonic kidney cyst model. In neonatal kidney-specific Pkd1 knockout mice, 1-Indanone remarkably slowed down kidney enlargement and cyst expansion. Furthermore, we demonstrated that 1-Indanone inhibited the abnormal elongation of cystic epithelial cilia by promoting tubulin polymerization and significantly down-regulating expression of anterograde transport motor protein KIF3A and IFT88. Moreover, we found that 1-Indanone significantly down-regulated ciliary coordinated Wnt/β-catenin, Hedgehog signaling pathways. These results demonstrate that 1-Indanone inhibits cystic cell proliferation by reducing abnormally prolonged cilia length in cystic epithelial cells, suggesting that 1-Indanone may hold therapeutic potential to retard cyst development in ADPKD.

Keywords:  1-Indanone; ADPKD; Cysts; anterograde transport motor protein; primary cilia

DOI:  https://doi.org/10.1038/s41401-022-00937-z
Nephron. 2022 Jul 27. 1-9

The Pathophysiology of Left Ventricular Hypertrophy, beyond Hypertension, in Autosomal Dominant Polycystic Kidney Disease.

Ozgur A Oto, Charles L Edelstein.

  Heart disease is one of the leading causes of death in autosomal dominant polycystic kidney disease (ADPKD) patients. Left ventricular hypertrophy (LVH) is an early and severe complication in ADPKD patients. Two decades ago, the prevalence of LVH on echocardiography in hypertensive ADPKD patients was shown to be as high as 46%. Recent studies using cardiac magnetic resonance imaging have shown that the prevalence of LVH in ADPKD patients may be lower. The true prevalence of LVH in ADPKD patients is controversial. There is evidence that factors other than hypertension contribute to LVH in ADPKD patients. Studies have shown that young normotensive ADPKD adults and children have a higher left ventricular mass index compared to controls and that the prevalence of LVH is high in patients with ADPKD whose blood pressure is well controlled. Polycystin-1 (PC-1) and polycystin-2 (PC-2) control intracellular signaling pathways that can influence cardiac function. Perturbations of PC-1 or PC-2 in the heart can lead to profound changes in cardiac structure and function independently of kidney function or blood pressure. PC-1 can influence mammalian target of rapamycin and mitophagy and PC-2 can influence autophagy, processes that play a role in LVH. Polymorphisms in the angiotensin-converting enzyme gene may play a role in LVH in ADPKD. This review will detail the pathophysiology of LVH, beyond hypertension, in ADPKD.

Keywords:  Cardiovascular disease; Left ventricular hypertrophy; Polycystic kidney disease

DOI:  https://doi.org/10.1159/000525944
Int J Mol Sci. 2022 Jul 23. pii: 8121. [Epub ahead of print]23(15):

Modulation of Primary Cilia by Alvocidib Inhibition of CILK1.

Elena X Wang, Jacob S Turner, David L Brautigan, Zheng Fu.

  The primary cilium provides cell sensory and signaling functions. Cilia structure and function are regulated by ciliogenesis-associated kinase 1 (CILK1). Ciliopathies caused by CILK1 mutations show longer cilia and abnormal Hedgehog signaling. Our study aimed to identify small molecular inhibitors of CILK1 that would enable pharmacological modulation of primary cilia. A previous screen of a chemical library for interactions with protein kinases revealed that Alvocidib has a picomolar binding affinity for CILK1. In this study, we show that Alvocidib potently inhibits CILK1 (IC50 = 20 nM), exhibits selectivity for inhibition of CILK1 over cyclin-dependent kinases 2/4/6 at low nanomolar concentrations, and induces CILK1-dependent cilia elongation. Our results support the use of Alvocidib to potently and selectively inhibit CILK1 to modulate primary cilia.

Keywords:  Alvocidib; CILK1; ICK; ciliopathy; kinase inhibitor; primary cilia

DOI:  https://doi.org/10.3390/ijms23158121
Hum Mol Genet. 2022 Jul 29. pii: ddac175. [Epub ahead of print]

Non-cell-autonomous activation of Hedgehog signaling contributes to disease progression in a mouse model of renal cystic ciliopathy.

Chia-Ling Hsieh, Stephanie Justine Jerman, Zhaoxia Sun.

Polycystic kidney disease (PKD) is a ciliopathy characterized by fluid-filled epithelial cysts in the kidney. Although it is well established that the primary cilium is essential for Hedgehog (HH) signalling and HH signalling is abnormally activated in multiple PKD models, the mechanism and function of HH activation in PKD pathogenesis remains incompletely understood. Here we used a transgenic HH reporter mouse line to identify the target tissue of HH signalling in Arl13f/f;Ksp-Cre mutant kidney, in which the cilia biogenesis gene Arl13b is specifically deleted in epithelial cells of the distal nephron. In addition, we used a co-culture system to dissect cross-talk between epithelial and mesenchymal cells in the absence of expanding cysts. Finally, we treated Arl13bf/f; Ksp-Cre mice with the GLI inhibitor GANT61 and analyzed its impact on PKD progression in this model. We found that deletion of Arl13b in epithelial cells in the mouse kidney, in vivo, led to non-cell-autonomous activation of the HH pathway in the interstitium. In vitro, when co-cultured with mesenchymal cells, Arl13b-/- epithelial cells produced more Sonic Hedgehog (SHH) in comparison to cells expressing Arl13b. Reciprocally, HH signalling was activated in mesenchymal cells co-cultured with Arl13b-/- epithelial cells. Finally, whole body inhibition of the HH pathway by GANT61 reduced the number of proliferating cells, inhibited cyst progression and fibrosis, and preserved kidney function in Arl13bf/f; Ksp-Cre mice. Our results reveal non-cell-autonomous activation of HH signalling in the interstitium of the Arl13bf/f; Ksp-Cre kidney and suggest that abnormal activation of the HH pathway contributes to disease progression.

DOI: https://doi.org/10.1093/hmg/ddac175
Am J Hum Genet. 2022 Jul 20. pii: S0002-9297(22)00266-X. [Epub ahead of print]

Monoallelic pathogenic ALG5 variants cause atypical polycystic kidney disease and interstitial fibrosis.

Genomics England Research Consortium

  Disorders of the autosomal dominant polycystic kidney disease (ADPKD) spectrum are characterized by the development of kidney cysts and progressive kidney function decline. PKD1 and PKD2, encoding polycystin (PC)1 and 2, are the two major genes associated with ADPKD; other genes include IFT140, GANAB, DNAJB11, and ALG9. Genetic testing remains inconclusive in ∼7% of the families. We performed whole-exome sequencing in a large multiplex genetically unresolved (GUR) family affected by ADPKD-like symptoms and identified a monoallelic frameshift variant (c.703_704delCA) in ALG5. ALG5 encodes an endoplasmic-reticulum-resident enzyme required for addition of glucose molecules to the assembling N-glycan precursors. To identify additional families, we screened a cohort of 1,213 families with ADPKD-like and/or autosomal-dominant tubulointerstitial kidney diseases (ADTKD), GUR (n = 137) or naive to genetic testing (n = 1,076), by targeted massively parallel sequencing, and we accessed Genomics England 100,000 Genomes Project data. Four additional families with pathogenic variants in ALG5 were identified. Clinical presentation was consistent in the 23 affected members, with non-enlarged cystic kidneys and few or no liver cysts; 8 subjects reached end-stage kidney disease from 62 to 91 years of age. We demonstrate that ALG5 haploinsufficiency is sufficient to alter the synthesis of the N-glycan chain in renal epithelial cells. We also show that ALG5 is required for PC1 maturation and membrane and ciliary localization and that heterozygous loss of ALG5 affects PC1 maturation. Overall, our results indicate that monoallelic variants of ALG5 lead to a disorder of the ADPKD-spectrum characterized by multiple small kidney cysts, progressive interstitial fibrosis, and kidney function decline.

Keywords:  ALG5; N-linked glycosylation; autosomal dominant tubulo-interstitial kidney disease; autosomal-dominant polycystic kidney disease; renal insufficiency

DOI:  https://doi.org/10.1016/j.ajhg.2022.06.013
Cells. 2022 Jul 22. pii: 2269. [Epub ahead of print]11(15):

A 3D In Vivo Model for Studying Human Renal Cystic Tissue and Mouse Kidney Slices.

Eva-Marie Bichlmayer, Lina Mahl, Leo Hesse, Eric Pion, Victoria Haller, Andreas Moehwald, Christina Hackl, Jens M Werner, Hans J Schlitt, Siegfried Schwarz, Philipp Kainz, Christoph Brochhausen, Christian Groeger, Felix Steger, Oliver Kölbl, Christoph Daniel, Kerstin Amann, Andre Kraus, Björn Buchholz, Thiha Aung, Silke Haerteis.

  (1) Background: Autosomal dominant polycystic kidney disease (ADPKD) is a frequent monogenic disorder that leads to progressive renal cyst growth and renal failure. Strategies to inhibit cyst growth in non-human cyst models have often failed in clinical trials. There is a significant need for models that enable studies of human cyst growth and drug trials. (2) Methods: Renal tissue from ADPKD patients who received a nephrectomy as well as adult mouse kidney slices were cultured on a chorioallantoic membrane (CAM) for one week. The cyst volume was monitored by microscopic and CT-based applications. The weight and angiogenesis were quantified. Morphometric and histological analyses were performed after the removal of the tissues from the CAM. (3) Results: The mouse and human renal tissue mostly remained vital for about one week on the CAM. The growth of cystic tissue was evaluated using microscopic and CT-based volume measurements, which correlated with weight and an increase in angiogenesis, and was accompanied by cyst cell proliferation. (4) Conclusions: The CAM model might bridge the gap between animal studies and clinical trials of human cyst growth, and provide a drug-testing platform for the inhibition of cyst enlargement. Real-time analyses of mouse kidney tissue may provide insights into renal physiology and reduce the need for animal experiments.

Keywords:  3D in vivo model; ADPKD; chorioallantoic membrane (CAM) model; human renal cystic tissue; mouse kidney slices; polycystic kidney disease

DOI:  https://doi.org/10.3390/cells11152269
J Am Soc Nephrol. 2022 Aug;33(8): 1433-1434

Polycystin-2 in the Endoplasmic Reticulum: Bending Ideas about the Role of the Cilium.

Michael J Caplan.



Keywords:  IP3 receptor; calcium; cilium; endoplasmic reticulum; polycystic kidney disease; polycystin-2

DOI:  https://doi.org/10.1681/ASN.2022050557
J Neurosci. 2022 Jul 25. pii: JN-RM-0497-22. [Epub ahead of print]

Disruption of dopamine receptor 1 localization to primary cilia impairs signaling in striatal neurons.

Toneisha Stubbs, Andrew Koemeter-Cox, James I Bingman, Fangli Zhao, Anuradha Kalyanasundaram, Leslie A Rowland, Muthu Periasamy, Calvin S Carter, Val C Sheffield, Candice C Askwith, Kirk Mykytyn.

A rod-shaped appendage called a primary cilium projects from the soma of most central neurons in the mammalian brain. The importance of cilia within the nervous system is highlighted by the fact that human syndromes linked to primary cilia dysfunction, collectively termed ciliopathies, are associated with numerous neuropathologies, including hyperphagia-induced obesity, neuropsychiatric disorders, and learning and memory deficits. Neuronal cilia are enriched with signaling molecules, including specific G protein-coupled receptors (GPCRs) and their downstream effectors, suggesting they act as sensory organelles that respond to neuromodulators in the extracellular space. We previously showed that GPCR ciliary localization is disrupted in neurons from mouse models of the ciliopathy Bardet-Biedl syndrome (BBS). Based on this finding we hypothesized that mislocalization of ciliary GPCRs may impact receptor signaling and contribute to the BBS phenotypes. Here, we show that disrupting localization of the ciliary GPCR dopamine receptor 1 (D1) in male and female mice, either by loss of a BBS protein or loss of the cilium itself, specifically in D1-expressing neurons, results in obesity. Interestingly, the weight gain is associated with reduced locomotor activity, rather than increased food intake. Moreover, loss of a BBS protein or cilia on D1-expressing neurons leads to a reduction in D1-mediated signaling. Together, these results indicate that cilia impact D1 activity in the nervous system and underscore the importance of neuronal cilia for proper GPCR signaling.SIGNIFICANCE STATEMENT:Most mammalian neurons possess solitary appendages called primary cilia. These rod-shaped structures are enriched with signaling proteins, such as G protein-coupled receptors (GPCRs), suggesting they respond to neuromodulators. This study examines the consequences of disrupting ciliary localization of the GPCR dopamine receptor 1 (D1) in D1-expressing neurons. Remarkably, mice that have either abnormal accumulation of D1 in cilia or loss of D1 ciliary localization become obese. In both cases the obesity is associated with lower locomotor activity rather than overeating. As D1 activation increases locomotor activity, these results are consistent with a reduction in D1 signaling. Indeed, we found that D1-mediated signaling is reduced in brain slices from both mouse models. Thus, cilia impact D1 signaling in the brain.

DOI: https://doi.org/10.1523/JNEUROSCI.0497-22.2022
Front Cell Dev Biol. 2022 ;10 901999

Manipulation of the Tubulin Code Alters Directional Cell Migration and Ciliogenesis.

Manuel Müller, Lena Gorek, Natalia Kamm, Ralf Jacob.

  Conjunction of epithelial cells into monolayer sheets implies the ability to migrate and to undergo apicobasal polarization. Both processes comprise reorganization of cytoskeletal elements and rearrangements of structural protein interactions. We modulated expression of tubulin tyrosin ligase (TTL), the enzyme that adds tyrosine to the carboxy terminus of detyrosinated α-tubulin, to study the role of tubulin detyrosination/-tyrosination in the orientation of cell motility and in epithelial morphogenesis. Oriented cell migration and the organization of focal adhesions significantly lose directionality with diminishing amounts of microtubules enriched in detyrosinated tubulin. On the other hand, increasing quantities of detyrosinated tubulin results in faster plus end elongation of microtubules in migrating and in polarized epithelial cells. These plus ends are decorated by the plus end binding protein 1 (EB1), which mediates interaction between microtubules enriched in detyrosinated tubulin and the integrin-ILK complex at focal adhesions. EB1 accumulates at the apical cell pole at the base of the primary cilium following apicobasal polarization. Polarized cells almost devoid of detyrosinated tubulin form stunted primary cilia and multiluminal cysts in 3D-matrices. We conclude that the balance between detyrosinated and tyrosinated tubulin alters microtubule dynamics, affects the orientation of focal adhesions and determines the organization of primary cilia on epithelial cells.

Keywords:  Focal Adhesion (FA); Tubulin Tyrosine Ligase (TTL); directional cell migration; epithelial morphogenesis; microtubule tyrosination/detyrosination; primary cilia

DOI:  https://doi.org/10.3389/fcell.2022.901999
Genes (Basel). 2022 Jul 01. pii: 1191. [Epub ahead of print]13(7):

TTC30A and TTC30B Redundancy Protects IFT Complex B Integrity and Its Pivotal Role in Ciliogenesis.

Felix Hoffmann, Sylvia Bolz, Katrin Junger, Franziska Klose, Timm Schubert, Franziska Woerz, Karsten Boldt, Marius Ueffing, Tina Beyer.

  Intraflagellar transport (IFT) is a microtubule-based system that supports the assembly and maintenance of cilia. The dysfunction of IFT leads to ciliopathies of variable severity. Two of the IFT-B components are the paralogue proteins TTC30A and TTC30B. To investigate whether these proteins constitute redundant functions, CRISPR/Cas9 was used to generate single TTC30A or B and double-knockout hTERT-RPE1 cells. Ciliogenesis assays showed the redundancy of both proteins while the polyglutamylation of cilia was affected in single knockouts. The localization of other IFT components was not affected by the depletion of a single paralogue. A loss of both proteins led to a severe ciliogenesis defect, resulting in no cilia formation, which was rescued by TTC30A or B. The redundancy can be explained by the highly similar interaction patterns of the paralogues; both equally interact with the IFT-B machinery. Our study demonstrates that a loss of one TTC30 paralogue can mostly be compensated by the other, thus preventing severe ciliary defects. However, cells assemble shorter cilia, which are potentially limited in their function, especially because of impaired polyglutamylation. A complete loss of both proteins leads to a deficit in IFT complex B integrity followed by disrupted IFT and subsequently no cilia formation.

Keywords:  CRISPR/Cas9; IFT; TTC30 paralogues; affinity proteomics; cilia; polyglutamylation

DOI:  https://doi.org/10.3390/genes13071191
Tomography. 2022 Jul 13. 8(4): 1804-1819

Deep Learning Automation of Kidney, Liver, and Spleen Segmentation for Organ Volume Measurements in Autosomal Dominant Polycystic Kidney Disease.

Arman Sharbatdaran, Dominick Romano, Kurt Teichman, Hreedi Dev, Syed I Raza, Akshay Goel, Mina C Moghadam, Jon D Blumenfeld, James M Chevalier, Daniil Shimonov, George Shih, Yi Wang, Martin R Prince.

  Organ volume measurements are a key metric for managing ADPKD (the most common inherited renal disease). However, measuring organ volumes is tedious and involves manually contouring organ outlines on multiple cross-sectional MRI or CT images. The automation of kidney contouring using deep learning has been proposed, as it has small errors compared to manual contouring. Here, a deployed open-source deep learning ADPKD kidney segmentation pipeline is extended to also measure liver and spleen volumes, which are also important. This 2D U-net deep learning approach was developed with radiologist labeled T2-weighted images from 215 ADPKD subjects (70% training = 151, 30% validation = 64). Additional ADPKD subjects were utilized for prospective (n = 30) and external (n = 30) validations for a total of 275 subjects. Image cropping previously optimized for kidneys was included in training but removed for the validation and inference to accommodate the liver which is closer to the image border. An effective algorithm was developed to adjudicate overlap voxels that are labeled as more than one organ. Left kidney, right kidney, liver and spleen labels had average errors of 3%, 7%, 3%, and 1%, respectively, on external validation and 5%, 6%, 5%, and 1% on prospective validation. Dice scores also showed that the deep learning model was close to the radiologist contouring, measuring 0.98, 0.96, 0.97 and 0.96 on external validation and 0.96, 0.96, 0.96 and 0.95 on prospective validation for left kidney, right kidney, liver and spleen, respectively. The time required for manual correction of deep learning segmentation errors was only 19:17 min compared to 33:04 min for manual segmentations, a 42% time saving (p = 0.004). Standard deviation of model assisted segmentations was reduced to 7, 5, 11, 5 mL for right kidney, left kidney, liver and spleen respectively from 14, 10, 55 and 14 mL for manual segmentations. Thus, deep learning reduces the radiologist time required to perform multiorgan segmentations in ADPKD and reduces measurement variability.

Keywords:  ADPKD; artificial intelligence; interobserver variability; kidney volume; liver volume; machine learning; spleen volume

DOI:  https://doi.org/10.3390/tomography8040152
Reproduction. 2022 Jul 01. pii: REP-21-0478. [Epub ahead of print]

Trophectoderm formation: regulation of morphogenesis and gene expressions by RHO, ROCK, cell polarity, and HIPPO signaling.

Vernadeth Alarcon, Yusuke Marikawa.

The trophectoderm (TE) is the first tissue to differentiate during the preimplantation development of placental mammals. It constitutes the outer epithelial wall of the blastocyst, and is responsible for hatching, uterine attachment, and placentation. Thus, its formation is the key initial step that enables the viviparity of mammals. Here, we first describe the general features of TE formation at the morphological and molecular levels. Prospective TE cells form an epithelial layer enclosing an expanding fluid-filled cavity by establishing the apical-basal cell polarity, intercellular junctions, microlumen, and osmotic gradient. A unique set of genes are expressed in TE that encode the transcription factors essential for the development of trophoblasts of the placenta upon implantation. TE-specific gene expressions are driven by the inhibition of HIPPO signaling, which is dependent on the prior establishment of the apical-basal polarity. We then discuss the specific roles of Ras homolog family members (RHO) and RHO-associated coiled-coil containing protein kinases (ROCK) as essential regulators of TE formation. RHO and ROCK modulate the actomyosin cytoskeleton, apical-basal polarity, intercellular junctions, and HIPPO signaling, thereby orchestrating the epithelialization and gene expressions in TE. Knowledge of the molecular mechanisms underlying TE formation is crucial for assisted reproductive technologies in human and farm animals, as it provides foundation to help improve procedures for embryo handling and selection to achieve better reproductive outcomes.

DOI: https://doi.org/10.1530/REP-21-0478
J Cell Sci. 2022 Jul 28. pii: jcs.260299. [Epub ahead of print]

Arih2 regulates hedgehog signaling through smoothened ubiquitination and ER-associated degradation.

Bo Lv, Xiao-Ou Zhang, Gregory J Pazour.

  During Hedgehog signaling, the ciliary levels of Ptch1 and Smo are regulated by the pathway. At the basal state, Ptch1 localizes to cilia and prevents the ciliary accumulation and activation of Smo. Upon binding Hedgehog ligand Ptch1 exits cilia, relieving inhibition of Smo. Smo then concentrates in cilia, becomes activated and activates downstream signaling. Loss of the ubiquitin E3 ligase Arih2 elevates basal Hedgehog signaling, elevates the cellular level of Smo, and increases basal levels of ciliary Smo. Mice express two isoforms of Arih2 with Arih2α found primarily in the nucleus and Arih2β found on the cytoplasmic face of the endoplasmic reticulum (ER). Re-expression of ER-localized Arih2β but not nuclear-localized Arih2α rescues the Hedgehog phenotypes. When Arih2 is defective, protein aggregates accumulate in the ER and the unfolded protein response is activated. Arih2β appears to regulate the ER-associated degradation of Smo preventing excess and potentially misfolded Smo from reaching the cilium and interfering with pathway regulation.

Keywords:  Arih2; Cilia; Hedgehog signaling; Ubiquitin

DOI:  https://doi.org/10.1242/jcs.260299
Proc Natl Acad Sci U S A. 2022 Aug 02. 119(31): e2201096119

Ciliogenesis requires sphingolipid-dependent membrane and axoneme interaction.

Dou Wu, Jingying Huang, Hao Zhu, Zhe Chen, Yongping Chai, Jingyi Ke, Kexin Lei, Zhao Peng, Ranhao Zhang, Xueming Li, Kaiyao Huang, Wei Li, Chengtian Zhao, Guangshuo Ou.

  Cilium formation and regeneration requires new protein synthesis, but the underlying cytosolic translational reprogramming remains largely unknown. Using ribosome footprinting, we performed global translatome profiling during cilia regeneration in Chlamydomonas and uncovered that flagellar genes undergo an early transcriptional activation but late translational repression. This pattern guided our identification of sphingolipid metabolism enzymes, including serine palmitoyltransferase (SPT), as essential regulators for ciliogenesis. Cryo-electron tomography showed that ceramide loss abnormally increased the membrane-axoneme distance and generated bulged cilia. We found that ceramides interact with intraflagellar transport (IFT) particle proteins that IFT motors transport along axoneme microtubules (MTs), suggesting that ceramide-IFT particle-IFT motor-MT interactions connect the ciliary membrane with the axoneme to form rod-shaped cilia. SPT-deficient vertebrate cells were defective in ciliogenesis, and SPT mutations from patients with hereditary sensory neuropathy disrupted cilia, which could be restored by sphingolipid supplementation. These results reveal a conserved role of sphingolipid in cilium formation and link compromised sphingolipid production with ciliopathies.

Keywords:  cilia; cryo-electron tomography; ribosome profiling; sphingolipid

DOI:  https://doi.org/10.1073/pnas.2201096119
Ther Adv Chronic Dis. 2022 ;13 20406223221112563

Evidence of nonsurgical treatment for polycystic liver disease.

Jeong-Ju Yoo, Hye In Jo, Eun-Ae Jung, Jae Seung Lee, Sang Gyune Kim, Young Seok Kim, Beom Kyung Kim.

  Background: Polycystic liver disease (PCLD) is the most common extrarenal manifestation of polycystic kidney disease. There is an urgent need to assess the efficacy and safety of nonsurgical modalities to relieve symptoms and decrease the severity of PCLD. Herein, we aimed to evaluate the efficacy of the nonsurgical treatment of PCLD and the quality of life of affected patients.Methods: PubMed, Ovid, MEDLINE, EMBASE, and the Cochrane Library were searched for studies on the nonsurgical modalities, either medications or radiological intervention to manage PCLD. Treatment efficacy, adverse events (AEs), and patient quality of life were evaluated.
Results: In total, 27 studies involving 1037 patients were selected. After nonsurgical treatment, liver volume decreased by 259 ml/m [mean change (Δ) of 6.22%] and the effect was higher in the radiological intervention group [-1617 ml/m (-15.49%)] than in the medication group [-151 ml/m (-3.78%)]. The AEs and serious AEs rates after overall nonsurgical treatment were 0.50 [95% confidence interval (CI): 0.33-0.67] and 0.04 (95% CI: 0.01-0.07), respectively. The results of the SF-36 questionnaire showed that PCLD treatment improved physical function [physical component summary score of 4.18 (95% CI: 1.54-6.83)] but did not significantly improve mental function [mental component summary score of 0.91 (95% CI: -1.20 to 3.03)].
Conclusion: Nonsurgical treatment was effective and safe for PCLD, but did not improve the quality of life in terms of mental health. Radiological intervention directly reduces hepatic cysts, and thus they should be considered for immediate symptom relief in patients with severe symptoms, whereas medication might be considered for maintenance treatment.
Registration number: PROSPERO (International Prospective Register of Systematic Reviews) CRD42021279597.

Keywords:  efficacy; intervention; medication; meta-analysis; polycystic liver disease; safety

DOI:  https://doi.org/10.1177/20406223221112563
Proc Natl Acad Sci U S A. 2022 Aug 02. 119(31): e2206098119

Regulated processing and secretion of a peptide precursor in cilia.

Raj Luxmi, Richard E Mains, Betty A Eipper, Stephen M King.

  Cilia are sensory and secretory organelles that both receive information from the environment and transmit signals. Cilia-derived vesicles (ectosomes), formed by outward budding of the ciliary membrane, carry enzymes and other bioactive products; this process represents an ancient mode of regulated secretion. Peptidergic intercellular communication controls a wide range of physiological and behavioral responses and occurs throughout eukaryotes. The Chlamydomonas reinhardtii genome encodes what appear to be numerous prepropeptides and enzymes homologous to those used to convert metazoan prepropeptides into bioactive peptide products. Since C. reinhardtii, a green alga, lack the dense core vesicles in which metazoan peptides are processed and stored, we explored the hypothesis that propeptide processing and secretion occur through the regulated release of ciliary ectosomes. A synthetic peptide (GATI-amide) that could be generated from a 91-kDa peptide precursor (proGATI) serves as a chemotactic modulator, attracting minus gametes while repelling plus gametes. Here we dissect the processing pathway that leads to formation of an amidated peptidergic sexual signal specifically on the ciliary ectosomes of plus gametes. Unlike metazoan propeptides, modeling studies identified stable domains in proGATI. Mass spectrometric analysis of a potential prohormone convertase and the amidated proGATI-derived products found in cilia and mating ectosomes link endoproteolytic cleavage to ectosome entry. Extensive posttranslational modification of proGATI confers stability to its amidated product. Analysis of this pathway affords insight into the evolution of peptidergic signaling; this will facilitate studies of the secretory functions of metazoan cilia.

Keywords:  Chlamydomonas; amidation; chemotaxis; ectosomes; peptidergic signaling

DOI:  https://doi.org/10.1073/pnas.2206098119
Front Genet. 2022 ;13 943197

Strongly Truncated Dnaaf4 Plays a Conserved Role in Drosophila Ciliary Dynein Assembly as Part of an R2TP-Like Co-Chaperone Complex With Dnaaf6.

Jennifer Lennon, Petra Zur Lage, Alex von Kriegsheim, Andrew P Jarman.

  Axonemal dynein motors are large multi-subunit complexes that drive ciliary movement. Cytoplasmic assembly of these motor complexes involves several co-chaperones, some of which are related to the R2TP co-chaperone complex. Mutations of these genes in humans cause the motile ciliopathy, Primary Ciliary Dyskinesia (PCD), but their different roles are not completely known. Two such dynein (axonemal) assembly factors (DNAAFs) that are thought to function together in an R2TP-like complex are DNAAF4 (DYX1C1) and DNAAF6 (PIH1D3). Here we investigate the Drosophila homologues, CG14921/Dnaaf4 and CG5048/Dnaaf6. Surprisingly, Drosophila Dnaaf4 is truncated such that it completely lacks a TPR domain, which in human DNAAF4 is likely required to recruit HSP90. Despite this, we provide evidence that Drosophila Dnaaf4 and Dnaaf6 proteins can associate in an R2TP-like complex that has a conserved role in dynein assembly. Both are specifically expressed and required during the development of the two Drosophila cell types with motile cilia: mechanosensory chordotonal neurons and sperm. Flies that lack Dnaaf4 or Dnaaf6 genes are viable but with impaired chordotonal neuron function and lack motile sperm. We provide molecular evidence that Dnaaf4 and Dnaaf6 are required for assembly of outer dynein arms (ODAs) and a subset of inner dynein arms (IDAs).

Keywords:  Drosophila; chaperone; ciliopathies; cilium; dynein; flagellum

DOI:  https://doi.org/10.3389/fgene.2022.943197
Int J Mol Sci. 2022 Jul 13. pii: 7749. [Epub ahead of print]23(14):

miR449 Protects Airway Regeneration by Controlling AURKA/HDAC6-Mediated Ciliary Disassembly.

Merit Wildung, Christian Herr, Dietmar Riedel, Cornelia Wiedwald, Alena Moiseenko, Fidel Ramírez, Hataitip Tasena, Maren Heimerl, Mihai Alevra, Naira Movsisyan, Maike Schuldt, Larisa Volceanov-Hahn, Sharen Provoost, Tabea Nöthe-Menchen, Diana Urrego, Bernard Freytag, Julia Wallmeier, Christoph Beisswenger, Robert Bals, Maarten van den Berge, Wim Timens, Pieter S Hiemstra, Corry-Anke Brandsma, Tania Maes, Stefan Andreas, Irene H Heijink, Luis A Pardo, Muriel Lizé.

  Airway mucociliary regeneration and function are key players for airway defense and are impaired in chronic obstructive pulmonary disease (COPD). Using transcriptome analysis in COPD-derived bronchial biopsies, we observed a positive correlation between cilia-related genes and microRNA-449 (miR449). In vitro, miR449 was strongly increased during airway epithelial mucociliary differentiation. In vivo, miR449 was upregulated during recovery from chemical or infective insults. miR0449-/- mice (both alleles are deleted) showed impaired ciliated epithelial regeneration after naphthalene and Haemophilus influenzae exposure, accompanied by more intense inflammation and emphysematous manifestations of COPD. The latter occurred spontaneously in aged miR449-/- mice. We identified Aurora kinase A and its effector target HDAC6 as key mediators in miR449-regulated ciliary homeostasis and epithelial regeneration. Aurora kinase A is downregulated upon miR449 overexpression in vitro and upregulated in miR449-/- mouse lungs. Accordingly, imaging studies showed profoundly altered cilia length and morphology accompanied by reduced mucociliary clearance. Pharmacological inhibition of HDAC6 rescued cilia length and coverage in miR449-/- cells, consistent with its tubulin-deacetylating function. Altogether, our study establishes a link between miR449, ciliary dysfunction, and COPD pathogenesis.

Keywords:  CDC20B; COPD; airway; ciliogenesis; lung; miR449; miR449a; microRNA

DOI:  https://doi.org/10.3390/ijms23147749
Int J Mol Sci. 2022 Jul 08. pii: 7582. [Epub ahead of print]23(14):

Retinal Ciliopathy in the Patient with Transplanted Kidney: Case Report.

Ivona Bućan, Mirjana Bjeloš, Irena Marković, Diana Bućan.

  A review of a rare case of a proven mutation in the RP1 gene (RP1c.2029C>T, p. (ARG677*) in a kidney transplant patient was presented herein. According to his medical history, he had tonsillectomy performed at the age of 20 due to erythrocyturia, and at the age of 32 he was treated for malignant hypertension. The patient had been diagnosed with chronic renal failure at age 56 years. During an eye examination in 2016, retinitis pigmentosa was suspected and the patient was advised to run further tests. After an ophthalmological examination and tests, genetic testing was performed and a mutation in the RP1 gene encoding a family of proteins which are components of microtubules in photoreceptor primary cilia was proven. The literature search found that mutations in the RP1 gene have so far been exclusively associated with a non-syndromic form of retinal degeneration. However, the RP1 protein is expressed in the kidneys, and it remains unclear why the mutation of this gene so far was only specifically related to retinal photoreceptor function and not to arterial hypertension and renal disease. Primary cilia are thought to act as potential mechanosensory fluid-flow receptors in the vascular endothelium and kidney and their dysfunction results in atherosclerotic changes, hypertension, and chronic renal failure.

Keywords:  RP1 gene; retinal–renal ciliopathies; retinitis pigmentosa

DOI:  https://doi.org/10.3390/ijms23147582
Front Genet. 2022 ;13 931822

Identical IFT140 Variants Cause Variable Skeletal Ciliopathy Phenotypes-Challenges for the Accurate Diagnosis.

Joanna Walczak-Sztulpa, Anna Wawrocka, Cenna Doornbos, Ronald van Beek, Anna Sowińska-Seidler, Aleksander Jamsheer, Ewelina Bukowska-Olech, Anna Latos-Bieleńska, Ryszard Grenda, Ernie M H F Bongers, Miriam Schmidts, Ewa Obersztyn, Maciej R Krawczyński, Machteld M Oud.

  Ciliopathies are rare congenital disorders, caused by defects in the cilium, that cover a broad clinical spectrum. A subgroup of ciliopathies showing significant phenotypic overlap are known as skeletal ciliopathies and include Jeune asphyxiating thoracic dysplasia (JATD), Mainzer-Saldino syndrome (MZSDS), cranioectodermal dysplasia (CED), and short-rib polydactyly (SRP). Ciliopathies are heterogeneous disorders with >187 associated genes, of which some genes are described to cause more than one ciliopathy phenotype. Both the clinical and molecular overlap make accurate diagnosing of these disorders challenging. We describe two unrelated Polish patients presenting with a skeletal ciliopathy who share the same compound heterozygous variants in IFT140 (NM_014,714.4) r.2765_2768del; p.(Tyr923Leufs*28) and exon 27-30 duplication; p.(Tyr1152_Thr1394dup). Apart from overlapping clinical symptoms the patients also show phenotypic differences; patient 1 showed more resemblance to a Mainzer-Saldino syndrome (MZSDS) phenotype, while patient 2 was more similar to the phenotype of cranioectodermal dysplasia (CED). In addition, functional testing in patient-derived fibroblasts revealed a distinct cilium phenotyps for each patient, and strikingly, the cilium phenotype of CED-like patient 2 resembled that of known CED patients. Besides two variants in IFT140, in depth exome analysis of ciliopathy associated genes revealed a likely-pathogenic heterozygous variant in INTU for patient 2 that possibly affects the same IFT-A complex to which IFT140 belongs and thereby could add to the phenotype of patient 2. Taken together, by combining genetic data, functional test results, and clinical findings we were able to accurately diagnose patient 1 with "IFT140-related ciliopathy with MZSDS-like features" and patient 2 with "IFT140-related ciliopathy with CED-like features". This study emphasizes that identical variants in one ciliopathy associated gene can lead to a variable ciliopathy phenotype and that an in depth and integrated analysis of clinical, molecular and functional data is necessary to accurately diagnose ciliopathy patients.

Keywords:  CED-like features; IFT140; MZSDS-like features; cilium phenotype; skeletal ciliopathy

DOI:  https://doi.org/10.3389/fgene.2022.931822
Genes (Basel). 2022 Jul 15. pii: 1252. [Epub ahead of print]13(7):

Challenges in Diagnosing Primary Ciliary Dyskinesia in a Brazilian Tertiary Hospital.

Mariana Dalbo Contrera Toro, José Dirceu Ribeiro, Fernando Augusto Lima Marson, Érica Ortiz, Adyléia Aparecida Dalbo Contrera Toro, Carmen Silvia Bertuzzo, Marcus Herbert Jones, Eulália Sakano.

  Primary ciliary dyskinesia (PCD) causes cellular cilia motility alterations, leading to clinical manifestations in the upper and lower respiratory tract and situs abnormalities. The PCD diagnosis was improved after the inclusion of diagnostic tools, such as transmission electron microscopy and genetic screening; however, the PCD screening is a challenge yet. In this context, we aimed to describe the clinical, genetic, and ultra-ciliary characteristics in individuals with clinical suspicion of PCD (cPCD) from a Brazilian Tertiary Hospital. An observational study was carried out with individuals during the follow-up between 2011 and 2021. The individuals were submitted to clinical questionnaires, transmission electron microscopy, and genetic screening for pathogenic variants in PCD-related genes. Those patients were classified according to the degree of suspicion for PCD. In our study, we enrolled thirty-seven cPCD individuals; 20/37 (54.1%) had chronic rhinosinusitis, 28/37 (75.6%) had bronchiectasis, and 29/37 (78.4%) had recurrent pneumonia. A total of 17/37 (45.9%) individuals had transmission electron microscopy or genetic confirmation of PCD; 10 individuals had at least one positive pathogenic genetic variant in the PCD-related genes; however, only seven patients presented a conclusive result according to the American College of Medical Genetics and Genomics and the Association for Molecular Pathology with two pathogenic variants in homozygous or compound heterozygous. The median age at diagnosis was 13 years, and the median time between suspicion and diagnosis was four years. Sixteen patients had class I electron microscopy alterations, seven had class II alterations, and 14 had normal transmission electron microscopy according to the international consensus guideline for reporting transmission electron microscopy results in the diagnosis of PCD (BEAT-PCD TEM Criteria). Genetic screening for pathogenic variants in PCD-related genes and transmission electron microscopy can help determine the PCD diagnosis; however, they are still unavailable to all individuals with clinical suspicion in Brazil. We described ultrastructural alterations found in our population along with the identification of pathogenic variants in PCD-related genes.

Keywords:  Kartagener syndrome; bronchiectasis; ciliary motility disorders; genetic testing; microscopy; sinusitis; transmission electron microscopy

DOI:  https://doi.org/10.3390/genes13071252