bims-bicyki 2021-12-12 papers

bims-bicyki

Biomed News

on Bicaudal-C1 and interactors in cystic kidney disease

Issue of 2021–12–12
eightteen papers selected by
Céline Gagnieux, École Polytechnique Fédérale de Lausanne

Antioxidant enzyme peroxiredoxin 5 regulates cyst growth and ciliogenesis via modulating Plk1 stability.
Monoallelic IFT140 pathogenic variants are an important cause of the autosomal dominant polycystic kidney-spectrum phenotype.
Translational research approaches to study pediatric polycystic kidney disease.
[Management of native kidneys in renal transplant recipients with autosomal dominant polycystic kidney].
Australian and New Zealand Scientific Meeting in Polycystic Kidney Disease (ANZ-PKD Meeting), Novotel Parramatta & Westmead Institute for Medical Research, Sydney, Australia, 11th -12th October, 2019.
Identification of ACOT13 and PTGER2 as novel candidate genes of autosomal dominant polycystic kidney disease through whole exome sequencing.
Assessing Risk of Progression in ADPKD.
Circ_HECTD1 regulates cerebral ischemia injury via mechanisms involving the regulation of let-7c-5p/ROCK1 axis.
Function of Centriolar Satellites and Regulation by Post-Translational Modifications.
Renal Neoplasia in Polycystic Kidney Disease: An Assessment of Tuberous Sclerosis Complex-associated Renal Neoplasia and PKD1/TSC2 Contiguous Gene Deletion Syndrome.
Ciliary Hedgehog signaling patterns the digestive system to generate mechanical forces driving elongation.
RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling.
The effects of somatostatin analogues on liver volume and quality of life in polycystic liver disease: a meta-analysis of randomized controlled trials.
Oligomerization of Ca2+/calmodulin-dependent protein kinase kinase.
Rab11a Regulates the Development of Cilia and Establishment of Planar Cell Polarity in Mammalian Vestibular Hair Cells.
Protocadherin-7 Regulates Osteoclast Differentiation through Intracellular SET-Binding Domain-Mediated RhoA and Rac1 Activation.
Licochalcone B specifically inhibits the NLRP3 inflammasome by disrupting NEK7-NLRP3 interaction.
The APC/C targets the Cep152-Cep63 complex at the centrosome to regulate mitotic spindle assembly.

FASEB J. 2022 Jan;36(1): e22089

Antioxidant enzyme peroxiredoxin 5 regulates cyst growth and ciliogenesis via modulating Plk1 stability.

Ewud Agborbesong, Julie Xia Zhou, Linda Xiaoyan Li, James P Calvet, Xiaogang Li.

  Oxidative stress is emerging as a contributing factor to the homeostasis in cystic diseases. However, the role antioxidant enzymes play in the pathogenesis of autosomal dominant polycystic kidney disease (ADPKD) remains elusive. Peroxiredoxin 5 (Prdx5) is an antioxidant enzyme that catalyzes the reduction of H2 O2 and alkyl hydroperoxide and plays an important role in different biological processes. In this study, we show that Prdx5 is downregulated in a PKD mutant mouse model and ADPKD patient kidneys. Knockdown of Prdx5 resulted in the formation of cysts in a three-dimensional mouse inner medullar collecting duct (IMCD) cell Matrigel culture system. The mechanisms of Prdx5 deficiency mediated cyst growth include: (1) induction of oxidative stress as indicated by increased mRNA expression of heme oxygenase-1, an oxidant stress marker; (2) activation of Erk, S6 and mTORC1, which contribute to cystic renal epithelial cell proliferation and cyst growth; (3) abnormal centrosome amplification and multipolar spindle formation which result in genome instability; (4) upregulation of Polo-like kinase 1 (Plk1) and Aurora kinase A, important mitotic kinases involved in cell proliferation and ciliogenesis; (5) impaired formation of primary cilia in mouse IMCD3 and retinal pigment epithelial cells, which could be rescued by inhibiting Plk1 activity; and (6) restraining the effect of Wnt3a and Wnt5a ligands on primary cilia in mouse IMCD3 cells, while regulating the activity of the canonical and non-canonical Wnt signaling in a separate cilia independent mechanism, respectively. Importantly, we found that targeting Plk1 with its inhibitor, volasertib, delayed cyst growth in Pkd1 conditional knockout mouse kidneys. Together, these findings indicate that Prdx5 is an important antioxidant that regulates cyst growth via diverse mechanisms, in particular, the Prdx5-Plk1 axis, and that induction and activation of Prdx5, alone or together with inhibition of Plk1, represent a promising strategy for combatting ADPKD.

Keywords:  ADPKD; Plk1; ciliogenesis; cyst growth; peroxiredoxin 5

DOI:  https://doi.org/10.1096/fj.202101270RR
Am J Hum Genet. 2021 Dec 07. pii: S0002-9297(21)00436-5. [Epub ahead of print]

Monoallelic IFT140 pathogenic variants are an important cause of the autosomal dominant polycystic kidney-spectrum phenotype.

Genomics England Research Consortium, the HALT PKD, CRISP, DIPAK, ADPKD Modifier, and TAME PKD studies

  Autosomal dominant polycystic kidney disease (ADPKD), characterized by progressive cyst formation/expansion, results in enlarged kidneys and often end stage kidney disease. ADPKD is genetically heterogeneous; PKD1 and PKD2 are the common loci (∼78% and ∼15% of families) and GANAB, DNAJB11, and ALG9 are minor genes. PKD is a ciliary-associated disease, a ciliopathy, and many syndromic ciliopathies have a PKD phenotype. In a multi-cohort/-site collaboration, we screened ADPKD-diagnosed families that were naive to genetic testing (n = 834) or for whom no PKD1 and PKD2 pathogenic variants had been identified (n = 381) with a PKD targeted next-generation sequencing panel (tNGS; n = 1,186) or whole-exome sequencing (WES; n = 29). We identified monoallelic IFT140 loss-of-function (LoF) variants in 12 multiplex families and 26 singletons (1.9% of naive families). IFT140 is a core component of the intraflagellar transport-complex A, responsible for retrograde ciliary trafficking and ciliary entry of membrane proteins; bi-allelic IFT140 variants cause the syndromic ciliopathy, short-rib thoracic dysplasia (SRTD9). The distinctive monoallelic phenotype is mild PKD with large cysts, limited kidney insufficiency, and few liver cysts. Analyses of the cystic kidney disease probands of Genomics England 100K showed that 2.1% had IFT140 LoF variants. Analysis of the UK Biobank cystic kidney disease group showed probands with IFT140 LoF variants as the third most common group, after PKD1 and PKD2. The proximity of IFT140 to PKD1 (∼0.5 Mb) in 16p13.3 can cause diagnostic confusion, and PKD1 variants could modify the IFT140 phenotype. Importantly, our studies link a ciliary structural protein to the ADPKD spectrum.

Keywords:  ADPKD; IFT140; cilia; ciliopathy; intraflagellar transport; monoallelic cystic disease; polycystic kidney disease; short rib thoracic dysplasia

DOI:  https://doi.org/10.1016/j.ajhg.2021.11.016
Mol Cell Pediatr. 2021 Dec 09. 8(1): 20

Translational research approaches to study pediatric polycystic kidney disease.

Max Christoph Liebau, Djalila Mekahli.

  Polycystic kidney diseases (PKD) are severe forms of genetic kidney disorders. The two main types of PKD are autosomal recessive and autosomal dominant PKD (ARPKD, ADPKD). While ARPKD typically is a disorder of early childhood, patients with ADPKD often remain pauci-symptomatic until adulthood even though formation of cysts in the kidney already begins in children. There is clinical and genetic overlap between both entities with very variable clinical courses. Subgroups of very early onset ADPKD may for example clinically resemble ARPKD. The basis of the clinical variability in both forms of PKD is not well understood and there are also limited prediction markers for disease progression for daily clinical life or surrogate endpoints for clinical trials in ARPKD or early ADPKD.As targeted therapeutic approaches to slow disease progression in PKD are emerging, it is becoming more important to reliably identify patients at risk for rapid progression as they might benefit from early therapy. Over the past years regional, national and international data collections to jointly analyze the clinical courses of PKD patients have been set up. The clinical observations are complemented by genetic studies and biorepositories as well as basic science approaches to elucidate the underlying molecular mechanisms in the PKD field. These approaches may serve as a basis for the development of novel therapeutic interventions in specific subgroups of patients. In this article we summarize some of the recent developments in the field with a focus on kidney involvement in PKD during childhood and adolescence and findings obtained in pediatric cohorts.

Keywords:  ADPedKD; ARegPKD; Ciliopathies; Genetic Kidney Disease; PKD1; PKD2; PKHD1

DOI:  https://doi.org/10.1186/s40348-021-00131-x
Rev Med Liege. 2021 Dec;76(12): 850-854

[Management of native kidneys in renal transplant recipients with autosomal dominant polycystic kidney].

A Thirion, M Sempels.

Autosomal dominant polycystic kidney disease (ADPKD) is responsible for 10 % of end-stage renal failure cases in Europe and the majority of these patients will have their renal failure treated with kidney transplantation. In this context, native kidneys will have a negligible function but maybe the source of a series of complications, whether due to polycystosis or immunosuppression. These complications include urinary tract infections, renal neoplasms, high blood pressure and abdominal pain and must be managed specifically for this particular context. Native nephrectomy may also be considered to prevent these complications, but it is a serious procedure and the risk-benefit ratio must be carefully assessed. There are still no clear and consensual guidelines on the indications for this nephrectomy or on the ideal timing of it in relation to the transplant procedure. Nevertheless, a review of the various data in the literature allows us to suggest an algorithm to help the therapeutic decision.

Keywords: Native nephrectomy; Polycystic kidney disease ; Renal transplantation ; ADPKD
Nephrology (Carlton). 2021 Dec;26 Suppl 3 7-19

Australian and New Zealand Scientific Meeting in Polycystic Kidney Disease (ANZ-PKD Meeting), Novotel Parramatta & Westmead Institute for Medical Research, Sydney, Australia, 11th -12th October, 2019.

DOI: https://doi.org/10.1111/nep.14001
Eur J Med Res. 2021 Dec 09. 26(1): 142

Identification of ACOT13 and PTGER2 as novel candidate genes of autosomal dominant polycystic kidney disease through whole exome sequencing.

Na Du, Dan Dong, Luyao Sun, Lihe Che, Xiaohua Li, Yong Liu, Bin Wang.

   BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is the most common monogenic kidney disorder. Half of the patients would slowly progress to end-stage renal disease. However, the potential target for ADPKD treatment is still lacking.
METHODS: Four ADPKD patients and two healthy family members were included in this study. The peripheral blood samples were obtained and tested by the whole exome sequencing (WES). The autosomal mutations in ADPKD patients were retained as candidate sites. The Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, and protein-protein interaction network (PPI) analyses were performed by clusterProfiler R package. A dataset containing 18 ADPKD patients and three normal samples were downloaded from the Gene Expression Omnibus (GEO) database and analyzed using the limma R package.
RESULTS: A total of six mutant genes were identified based on the dominant genetic pattern and most of them had not been reported to be associated with ADPKD. Furthermore, 19 harmful genes were selected according to the harmfulness of mutation. GO and KEGG enrichment analyses showed that the processes of single-organism cellular process, response to stimulus, plasma membrane, cell periphery, and anion binding as well as cyclic adenosine monophosphate (cAMP) signaling pathway and pathways in cancer were significantly enriched. Through integrating PPI and gene expression analyses, acyl-CoA thioesterase 13 (ACOT13), which has not been reported to be related to ADPKD, and prostaglandin E receptor 2 (PTGER2) were identified as potential genes associated with ADPKD.
CONCLUSIONS: Through combination of WES, gene expression, and PPI network analyses, we identified ACOT13 and PTGER2 as potential ADPKD-related genes.

Keywords:  ACOT13; Gene mutations; PTGER2; Polycystic kidney disease; Whole exome sequencing

DOI:  https://doi.org/10.1186/s40001-021-00613-8
Clin J Am Soc Nephrol. 2021 Dec 09. pii: CJN.13071021. [Epub ahead of print]

Assessing Risk of Progression in ADPKD.

Craig E Gordon, Dana C Miskulin, Ronald D Perrone.



Keywords:  autosomal dominant polycystic kidney disease; chronic kidney disease; clinical nephrology; genetic renal disease; polycystic kidney disease; randomized controlled trials

DOI:  https://doi.org/10.2215/CJN.13071021
Neuroreport. 2022 Jan 05. 33(1): 13-22

Circ_HECTD1 regulates cerebral ischemia injury via mechanisms involving the regulation of let-7c-5p/ROCK1 axis.

Qian Guo, Tingting Guo, Yuanyuan Sun, Tianrong Chen.

BACKGROUND: Ischemia is the main cause of cerebral ischemic stroke with a high mortality rate, and it is affected by the dysfunction of circular RNAs. The underlying molecular mechanisms of circ_HECTD1 were explored in cerebral ischemia stroke.
METHODS: PC-12 cells were treated with oxygen-glucose deprivation/reoxygenation (OGD/R) as the cell model of cerebral ischemia model. The expression levels of circ_HECTD1, let-7c-5p, and Rho-associated coiled-coil-containing protein kinase 1 (ROCK1) were determined by quantitative real-time PCR in PC-12 cells. The proliferation ability of PC-12 cells was assessed by 5-ethynyl-2'-deoxyuridine and 3-(4,5-dimethylthizol-2-yl)-2,5-diphenyltetrazolium bromide assays. The protein expression levels were quantified by western blot analysis. Flow cytometry was performed to analyze cell apoptosis. Lactate dehydrogenase concentration was assessed by a commercialized kit. Dual-luciferase reporter assay was used to confirm the interaction relationships among circ_HECTD1, let-7c-5p and ROCK1.
RESULTS: Circ_HECTD1 was upregulated in OGD/R-treated PC-12 cells. The results also showed that cell proliferation was decreased and apoptosis was increased in OGD/R-treated PC-12 cells, which was overturned by the inhibition of circ_HECTD1. Let-7c-5p was a target of circ_HECTD1, and the protective effects of circ_HECTD1 knockdown on OGD/R-treated PC-12 cells were canceled after co-transfection with let-7c-5p inhibitor. We found that ROCK1 was a potential target of let-7c-5p. Let-7c-5p -mediated the effects on the proliferation and apoptosis of OGD/R-treated PC-12 cells by targeting ROCK1.
CONCLUSION: Circ_HECTD1 was implicated in the development of cerebral ischemia stroke. Knockdown of circ_HECTD1 protected against cerebral ischemia injury in OGD/R-treated PC-12 cells depending on the regulation of let-7c-5p/ROCK1 axis.

DOI: https://doi.org/10.1097/WNR.0000000000001743
Front Cell Dev Biol. 2021 ;9 780502

Function of Centriolar Satellites and Regulation by Post-Translational Modifications.

Clotilde C N Renaud, Nicolas Bidère.

  Centriolar satellites are small membrane-less granules that gravitate around the centrosome. Recent advances in defining the satellite proteome and interactome have unveiled hundreds of new satellite components thus illustrating the complex nature of these particles. Although initially linked to the homeostasis of centrosome and the formation of primary cilia, these composite and highly dynamic structures appear to participate in additional cellular processes, such as proteostasis, autophagy, and cellular stress. In this review, we first outline the main features and many roles of centriolar satellites. We then discuss how post-translational modifications, such as phosphorylation and ubiquitination, shape their composition and functions. This is of particular interest as interfering with these processes may provide ways to manipulate these structures.

Keywords:  centriolar satellites; centrosome; phosphorylation; post-translational modification (PMT); primary cilia; ubiquitin

DOI:  https://doi.org/10.3389/fcell.2021.780502
Eur Urol. 2021 Dec 04. pii: S0302-2838(21)02161-8. [Epub ahead of print]

Renal Neoplasia in Polycystic Kidney Disease: An Assessment of Tuberous Sclerosis Complex-associated Renal Neoplasia and PKD1/TSC2 Contiguous Gene Deletion Syndrome.

Sounak Gupta, Christine M Lohse, Ross Rowsey, Michael R McCarthy, Wei Shen, Loren Herrera-Hernandez, Stephen A Boorjian, R Houston Thompson, Rafael E Jimenez, Bradley C Leibovich, John C Cheville.

  To determine the incidence of renal neoplasia among patients undergoing nephrectomy for polycystic kidney disease (PKD), we queried our institutional nephrectomy registry (years 2000-2020). Approximately 4% (231 of 5757) of patients who underwent nephrectomy had PKD, and 26 of these 231 patients (11.3%) had renal neoplasia. Tumors from an additional two patients with PKD were also evaluated. Patients with PKD who had tuberous sclerosis complex (TSC)-associated renal neoplasia were screened for PKD1/TSC2 contiguous gene deletion syndrome (CGS) using single nucleotide polymorphism arrays. The median age of patients with PKD and renal neoplasia at nephrectomy was 54 yr. The median tumor size was 2.0 cm and the tumors were predominantly of low grade and stage. The tumors consisted of 23 renal cell carcinomas (RCCs), one epithelioid angiomyolipoma, and four angiomyolipomas. The median follow-up was 59.5 mo (n = 26) and only one patient with clear cell RCC developed metastases. Two patients with angiomyolipomas had PKD1/TSC2 CGS. Our results support screening of patients with PKD and TSC-associated renal neoplasia as well as TSC patients with cystic renal disease for CGS, as identification of patients with CGS can better define the manifestation and prognosis of CGS and guide counseling regarding patterns of inheritance. PATIENT SUMMARY: We identified patients with abnormal kidney cell growth (called renal neoplasia) among those undergoing removal of kidney tissue for polycystic kidney disease (PKD) and patients with a syndrome involving deletions in two genes, called PKD1/TSC2 contiguous gene deletion syndrome (CGS) at our institution. Of 231 PKD patients with removal of kidney tissue, 11.3% had renal neoplasia, and two patients with angiomyolipoma tumors had PKD1/TSC2 CGS. Detection of renal neoplasia associated with a condition called tuberous sclerosis complex in PKD may increase the identification of patients with PKD1/TSC2 CGS and guide patient counseling regarding outcomes and patterns of inheritance.

Keywords:  PKD1/TSC2 contiguous gene deletion syndrome; Polycystic kidney disease; Tuberous sclerosis complex

DOI:  https://doi.org/10.1016/j.eururo.2021.11.013
Nat Commun. 2021 Dec 10. 12(1): 7186

Ciliary Hedgehog signaling patterns the digestive system to generate mechanical forces driving elongation.

Ying Yang, Pekka Paivinen, Chang Xie, Alexis Leigh Krup, Tomi P Makela, Keith E Mostov, Jeremy F Reiter.

How tubular organs elongate is poorly understood. We found that attenuated ciliary Hedgehog signaling in the gut wall impaired patterning of the circumferential smooth muscle and inhibited proliferation and elongation of developing intestine and esophagus. Similarly, ablation of gut-wall smooth muscle cells reduced lengthening. Disruption of ciliary Hedgehog signaling or removal of smooth muscle reduced residual stress within the gut wall and decreased activity of the mechanotransductive effector YAP. Removing YAP in the mesenchyme also reduced proliferation and elongation, but without affecting smooth muscle formation, suggesting that YAP interprets the smooth muscle-generated force to promote longitudinal growth. Additionally, we developed an intestinal culture system that recapitulates the requirements for cilia and mechanical forces in elongation. Pharmacologically activating YAP in this system restored elongation of cilia-deficient intestines. Thus, our results reveal that ciliary Hedgehog signaling patterns the circumferential smooth muscle to generate radial mechanical forces that activate YAP and elongate the gut.

DOI: https://doi.org/10.1038/s41467-021-27319-z
Redox Biol. 2021 Dec 06. pii: S2213-2317(21)00364-5. [Epub ahead of print]48 102204

RND3 attenuates oxidative stress and vascular remodeling in spontaneously hypertensive rat via inhibiting ROCK1 signaling.

Nan Wu, Fen Zheng, Na Li, Ying Han, Xiao-Qing Xiong, Jue-Jin Wang, Qi Chen, Yue-Hua Li, Guo-Qing Zhu, Ye-Bo Zhou.

  Superoxide and vascular smooth muscle cells (VSMCs) migration and proliferation play crucial roles in the vascular remodeling. Vascular remodeling contributes to the development and complications of hypertension. Rho family GTPase 3 (RND3 or RhoE), an atypical small Rho-GTPase, is known to be involved in cancer development and metastasis. However, the roles of RND3 in superoxide production and cardiovascular remodeling are unknown. Here, we uncovered the critical roles of RND3 in attenuating superoxide production, VSMCs migration and proliferation, and vascular remodeling in hypertension and its underline mechanisms. VSMCs were isolated and prepared from thoracic aorta of Male Wistar-Kyoto rat (WKY) and spontaneously hypertensive rat (SHR). RND3 mRNA and protein expressions in arteries and VSMCs were down-regulated in SHR. RND3 overexpression in VSMCs reduced NAD(P)H oxidase (NOX) activity, NOX1 and NOX2 expressions, mitochondria superoxide generation, and H2O2 production in SHR. Moreover, the RND3 overexpression inhibited VSMCs migration and proliferation in SHR, which were similar to the effects of NOX1 inhibitor ML171 plus NOX2 inhibitor GSK2795039. Rho-associated kinase 1 (ROCK1) and RhoA expressions and myosin phosphatase targeting protein 1 (MYPT1) phosphorylation in VSMCs were increased in SHR, which were prevented by RND3 overexpression. ROCK1 overexpression promoted NOX1 and NOX2 expressions, superoxide and H2O2 production, VSMCs migration and proliferation in both WKY and SHR, which were attenuated by RND3 overexpression. Adenoviral-mediated RND3 overexpression in SHR attenuated hypertension, vascular remodeling and oxidative stress. These results indicate that RND3 attenuates VSMCs migration and proliferation, hypertension and vascular remodeling in SHR via inhibiting ROCK1-NOX1/2 and mitochondria superoxide signaling.

Keywords:  Hypertension; Migration; Proliferation; RND3; Superoxide; Vascular remodeling; Vascular smooth muscle cells

DOI:  https://doi.org/10.1016/j.redox.2021.102204
Sci Rep. 2021 Dec 06. 11(1): 23500

The effects of somatostatin analogues on liver volume and quality of life in polycystic liver disease: a meta-analysis of randomized controlled trials.

Carlo Garofalo, Ivana Capuano, Luigi Pennino, Ilaria De Gregorio, Eleonora Riccio, Michele Provenzano, Felice Crocetto, Pasquale Buonanno, Savio Domenico Pandolfo, Michele Andreucci, Antonio Pisani.

A clear evidence on the benefits of somatostatin analogues (SA) on liver outcome in patients affected by polycystic liver disease is still lacking. We performed a meta-analysis of RCTs and a trial sequential analysis (TSA) evaluating the effects of SA in adult patients with polycystic liver disease on change in liver volume. As secondary outcome, we evaluated the effects on quality of life as measured by SF36-questionnaire. Six RCTs were selected with an overall sample size of 332 adult patients with polycystic liver disease (mean age: 46 years). Mean liver volume at baseline was 3289 ml in SA group and 3089 ml in placebo group. Overall, unstandardized mean difference in liver volume was - 176 ml (95%CI, - 406, 54; p < 0.133). Heterogeneity was low (I2:0%, p < 0.992). However, we performed a moderator analysis and we found that a higher eGFR significantly correlates to a more pronounced effect of SA on liver volume reduction (p = 0.036). Cumulative Z-curve in TSA did not reach either significance and futility boundaries or required information size. Three RCTs have evaluated Quality of life parameters measured by SF36-QOL questionnaire for a total of 124 patients; no significant difference was found on the effect of SA on QOL parameters when compared with placebo. The present meta-analysis revealed a potential effect of SA on reduction of liver volume and quality of life parameters, but results did not reach a statistical significance. These data could be explained by the need of further studies, as demonstrated through TSA, to reach an adequate sample size to confirm the beneficial outcomes of SAs treatment.

DOI: https://doi.org/10.1038/s41598-021-02812-z
Biochem Biophys Res Commun. 2021 Dec 01. pii: S0006-291X(21)01630-2. [Epub ahead of print]587 160-165

Oligomerization of Ca2+/calmodulin-dependent protein kinase kinase.

Yusei Fukumoto, Yuhei Harada, Satomi Ohtsuka, Naoki Kanayama, Masaki Magari, Naoya Hatano, Hiroyuki Sakagami, Hiroshi Tokumitsu.

  Ca2+/calmodulin-dependent protein kinase kinases (CaMKKα and β) are regulatory kinases for multiple downstream kinases, including CaMKI, CaMKIV, PKB/Akt, and AMP-activated protein kinase (AMPK) through phosphorylation of each activation-loop Thr residue. In this report, we biochemically characterize the oligomeric structure of CaMKK isoforms through a heterologous expression system using COS-7 cells. Oligomerization of CaMKK isoforms was readily observed by treating CaMKK transfected cells with cell membrane permeable crosslinkers. In addition, His-tagged CaMKKα (His-CaMKKα) pulled down with FLAG-tagged CaMKKα (FLAG-CaMKKα) in transfected cells. The oligomerization of CaMKKα was confirmed by the fact that GST-CaMKKα/His-CaMKKα complex from transiently expressed COS-7 cells extracts was purified to near homogeneity by the sequential chromatography using glutathione-sepharose/Ni-sepharose and was observed in a Ca2+/CaM-independent manner by reciprocal pulldown assay, suggesting the direct interaction between monomeric CaMKKα. Furthermore, the His-CaMKKα kinase-dead mutant (D293A) complexed with FLAG-CaMKKα exhibited significant CaMKK activity, indicating the active CaMKKα multimeric complex. Collectively, these results suggest that CaMKKα can self-associate in the cells, constituting a catalytically active oligomer that might be important for the efficient activation of CaMKK-mediated intracellular signaling.

Keywords:  Ca(2+)-signaling; CaM kinase cascade; CaMKK; Oligomerization; Phosphorylation

DOI:  https://doi.org/10.1016/j.bbrc.2021.11.105
Front Mol Neurosci. 2021 ;14 762916

Rab11a Regulates the Development of Cilia and Establishment of Planar Cell Polarity in Mammalian Vestibular Hair Cells.

Bin-Jun Chen, Xiao-Qing Qian, Xiao-Yu Yang, Tao Jiang, Yan-Mei Wang, Ji-Han Lyu, Fang-Lu Chi, Ping Chen, Dong-Dong Ren.

  Vestibular organs have unique planar cell polarity (Figure 1A), and their normal development and function are dependent on the regular polarity of cilia (Figure 1B) requires. Rab11a is a small G protein that participates in the transportation of intracellular and extracellular materials required for polarity formation; however, our understanding of the mechanisms of the actions of Rab11a in vestibular organs is limited. Here, we showed that the general shape of the utricle was abnormal in Rab11a CKO/CKO mice. These mice also showed abnormal morphology of the stereocilia bundles, which were reduced in both length and number, as well as disturbed tissue-level polarity. Rab11a affected the distribution of polarity proteins in the vestibular organs, indicating that the normal development of cilia requires Rab11a and intraflagellar transportation. Furthermore, small G protein migration works together with intraflagellar transportation in the normal development of cilia. FIGURE 1Morphological changes of stereocilia in the extrastriolar hair cells from Rab11a single or Rab11a/IFT88 double-mutant utricles. (A) Medial view of a mouse left inner ear with its five vestibular sensory organs (gray). Enlarged are the utricle showing their subdivisions, LPR (yellow line), and striola (blue). LES, lateral extrastriola; MES, medial extrastriola; LPR, line of polarity reversal. (B) Schematic view of vestibular hair cell. Kinocilium is marked with ace-tubulin. Basal body is marked with γ-tubulin. (C,C1,D,D1) Normal appearance of the stereocilia of extrastriolar hair cells of wild-type controls. (E,E1,F,F1) Altered morphology in Rab11a CKO/CKO animals. (G,G1,H,H1) The changes in the stereocilia morphology were more severe in Rab11a CKO/CKO /IFT 88 CKO/+ mice. (I-L) Higher magnification of confocal images of hair cells. (M-P) Scanning electron microscopy images of hair cells from wild-type controls and Rab11a mutants. (I,M) Morphology of normal. hair cells of wild-type controls. (J,N) The number of stereocilia on a single hair cell was deceased in the Rab11a mutant. (K,O) Stereocilia were shorter in mutants compared to the wild-type controls. (L,P) The staircase-like hair bundle architecture of hair cells was lost in Rab11a mutant mice. (Q) The percentage of hair cells with abnormal development of static cilia bundles in the extrastriola region was counted as a percentage of the total (n = 5). The percentage of abnormal hair cells was higher in Rab11a CKO/CKO , IFT88 CKO/+ mice compared to Rab11a CKO/CKO . The abnormal ratios of single and double knockout hair cells were 42.1 ± 5.7 and 71.5 ± 10.4, respectively. In (A-J), for all primary panels, hair cell stereociliary bundles were marked with phalloidin (green), the actin-rich cuticular plate of hair cells was labeled with β-spectrin (red), while the basal body of the hair cell was labeled with γ-tubulin (blue). Scale bars: 10 μm (C-H1), 5 μm (J-N). *P < 0.05.

Keywords:  Rab11a; cilia; development; planar cell polarity; vestibular

DOI:  https://doi.org/10.3389/fnmol.2021.762916
Int J Mol Sci. 2021 Dec 04. pii: 13117. [Epub ahead of print]22(23):

Protocadherin-7 Regulates Osteoclast Differentiation through Intracellular SET-Binding Domain-Mediated RhoA and Rac1 Activation.

Hyunsoo Kim, Noriko Takegahara, Yongwon Choi.

  Protocadherin-7 (Pcdh7) is a member of the non-clustered protocadherin δ1 subgroup of the cadherin superfamily. Although the cell-intrinsic role of Pcdh7 in osteoclast differentiation has been demonstrated, the molecular mechanisms of Pcdh7 regulating osteoclast differentiation remain to be determined. Here, we demonstrate that Pcdh7 contributes to osteoclast differentiation by regulating small GTPases, RhoA and Rac1, through its SET oncoprotein binding domain. Pcdh7 is associated with SET along with RhoA and Rac1 during osteoclast differentiation. Pcdh7-deficient (Pcdh7-/-) cells showed abolished RANKL-induced RhoA and Rac1 activation, and impaired osteoclast differentiation. Impaired osteoclast differentiation in Pcdh7-/- cells was restored by retroviral transduction of full-length Pcdh7 but not by a Pcdh7 mutant that lacks SET binding domain. The direct crosslink of the Pcdh7 intracellular region induced the activation of RhoA and Rac1, which was not observed when Pcdh7 lacks the SET binding domain. Additionally, retroviral transduction of the constitutively active form of RhoA and Rac1 completely restored the impaired osteoclast differentiation in Pcdh7-/- cells. Collectively, these results demonstrate that Pcdh7 controls osteoclast differentiation by regulating RhoA and Rac1 activation through the SET binding domain.

Keywords:  Pcdh7; Rac1; RhoA; SET; differentiation; osteoclast

DOI:  https://doi.org/10.3390/ijms222313117
EMBO Rep. 2021 Dec 09. e53499

Licochalcone B specifically inhibits the NLRP3 inflammasome by disrupting NEK7-NLRP3 interaction.

Qiang Li, Hui Feng, Hongbo Wang, Yinghao Wang, Wenqing Mou, Guang Xu, Ping Zhang, Ruisheng Li, Wei Shi, Zhilei Wang, Zhie Fang, Lutong Ren, Yan Wang, Li Lin, Xiaorong Hou, Wenzhang Dai, Zhiyong Li, Ziying Wei, Tingting Liu, Jiabo Wang, Yuming Guo, Pengyan Li, Xu Zhao, Xiaoyan Zhan, Xiaohe Xiao, Zhaofang Bai.

  The activation of the nucleotide oligomerization domain (NOD)-like receptor (NLR) family, pyrin domain-containing protein 3 (NLRP3) inflammasome is related to the pathogenesis of a wide range of inflammatory diseases, but drugs targeting the NLRP3 inflammasome are still scarce. In the present study, we demonstrated that Licochalcone B (LicoB), a main component of the traditional medicinal herb licorice, is a specific inhibitor of the NLRP3 inflammasome. LicoB inhibits the activation of the NLRP3 inflammasome in macrophages but has no effect on the activation of AIM2 or NLRC4 inflammasome. Mechanistically, LicoB directly binds to NEK7 and inhibits the interaction between NLRP3 and NEK7, thus suppressing NLRP3 inflammasome activation. Furthermore, LicoB exhibits protective effects in mouse models of NLRP3 inflammasome-mediated diseases, including lipopolysaccharide (LPS)-induced septic shock, MSU-induced peritonitis and non-alcoholic steatohepatitis (NASH). Our findings indicate that LicoB is a specific NLRP3 inhibitor and a promising candidate for treating NLRP3 inflammasome-related diseases.

Keywords:  LPS-induced septic shock; Licochalcone B; MSU-induced peritonitis; NASH; NEK7; NLRP3 inflammasome

DOI:  https://doi.org/10.15252/embr.202153499
J Cell Sci. 2021 Dec 08. pii: jcs.259273. [Epub ahead of print]

The APC/C targets the Cep152-Cep63 complex at the centrosome to regulate mitotic spindle assembly.

Thomas Tischer, Jing Yang, David Barford.

  The control of protein abundance is a fundamental regulatory mechanism during mitosis. The anaphase promoting complex/cyclosome (APC/C) is the main protein ubiquitin ligase responsible for the temporal regulation of mitotic progression. It has been proposed that the APC/C might fulfil other functions including assembly of the mitotic spindle. Here, we show that the APC/C localizes to centrosomes, the organizers of the eukaryotic microtubule cytoskeleton, specifically during mitosis. Recruitment of the APC/C to spindle poles requires the centrosomal protein Cep152, and we identified Cep152 as both an APC/C interaction partner and as an APC/C substrate. Previous studies showed that Cep152 forms a complex with Cep57 and Cep63. The APC/C-mediated ubiquitination of Cep152 at the centrosome releases Cep57 from this inhibitory complex and enables its interaction with pericentrin, a critical step in promoting microtubule nucleation. Thus, our study extends the function of the APC/C from being a regulator of mitosis to also acting as a positive governor of spindle assembly. The APC/C thereby integrates control of these two important processes in a temporal manner.

Keywords:  APC/C; Centrosome; Cep152; Microtubules; Mitosis; Ubiquitin

DOI:  https://doi.org/10.1242/jcs.259273