bims-tubesc Biomed News
on Molecular mechanisms in tuberous sclerosis
Issue of 2021‒05‒23
eleven papers selected by
Marti Cadena Sandoval

  1. Child Neurol Open. 2021 Jan-Dec;8:8 2329048X211012817
      Objective: We aimed to further elucidate the phenotypic spectrum of Tuberous Sclerosis Complex (TSC) depending on genotype.Methods: A retrospective review of patients seen in the TSC clinic at the Hospital for Sick Children was conducted and the frequency of TSC manifestations was compared based on genotype.
    Results: Nineteen-patients had TSC1 mutations, 36 had TSC2 mutations and 11 had no mutation identified (NMI). Patients with TSC2 mutations had a higher frequency of early-onset epilepsy and more frequent systemic manifestations. The NMI group had milder neurologic and systemic manifestations. Our data did not demonstrate that intellectual disability and infantile spasms were more common in TSC2 mutations.
    Conclusions: This is the first Canadian pediatric cohort exploring the genotype-phenotype relationship in TSC. We report that some manifestations are more frequent and severe in TSC2 mutations and that NMI may have a milder phenotype. Disease surveillance and counseling should continue regardless of genotype until this is better elucidated.
    Keywords:  genotype; no mutation identified (NMI); phenotype; surveillance; tuberous sclerosis complex (TSC)
  2. J Biol Chem. 2021 May 14. pii: S0021-9258(21)00573-1. [Epub ahead of print] 100780
      Macroautophagy (hereafter, autophagy) is a process that directs the degradation of cytoplasmic material in lysosomes. In addition to its homeostatic roles, autophagy undergoes dynamic positive and negative regulation in response to multiple forms of cellular stress, thus enabling the survival of cells. However, the precise mechanisms of autophagy regulation are not fully understood. To identify potential negative regulators of autophagy, we performed a genome-wide CRISPR screen using the quantitative autophagic flux reporter GFP-LC3-RFP. We identified phosphoribosylformylglycinamidine synthase (PFAS), a component of the de novo purine synthesis pathway, as one such negative regulator of autophagy. Autophagy was activated in cells lacking PFAS or phosphoribosyl pyrophosphate amidotransferase (PPAT), another de novo purine synthesis enzyme, or treated with methotrexate when exogenous levels of purines were insufficient. Purine starvation-induced autophagy activation was concomitant with mTORC1 suppression, and was profoundly suppressed in cells deficient for TSC2, which negatively regulates mTORC1 through inhibition of RHEB, suggesting that purines regulate autophagy through the TSC-RHEB-mTORC1 signaling axis. Moreover, depletion of the pyrimidine synthesis enzymes carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD) and dihydroorotate dehydrogenase (DHODH) activated autophagy as well, although mTORC1 activity was not altered by pyrimidine shortage. These results suggest a different mechanism of autophagy induction between purine and pyrimidine starvation. These findings provide novel insights into the regulation of autophagy by nucleotides and possibly the role of autophagy in nucleotide metabolism, leading to further developing anticancer strategies involving nucleotide synthesis and autophagy.
    Keywords:  CRISPR/Cas; mammalian target of rapamycin (mTOR); nucleoside/nucleotide biosynthesis; nucleoside/nucleotide metabolism; nucleotide; phosphoribosylformylglycinamidine synthase (PFAS); tuberous sclerosis complex (TSC)
  3. Cell Mol Biol Lett. 2021 May 18. 26(1): 18
      BACKGROUND: Mammalian/mechanistic target of rapamycin (mTOR) complexes are essential for cell proliferation, growth, differentiation, and survival. mTORC1 hyperactivation occurs in the tuberous sclerosis complex (TSC). mTORC1 localizes to the surface of lysosomes, where Rheb activates it. However, mTOR was also found on the endoplasmic reticulum (ER) and Golgi apparatus (GA). Recent studies showed that the same inputs regulate ER-to-GA cargo transport and mTORC1 (e.g., the level of amino acids or energy status of the cell). Nonetheless, it remains unknown whether mTOR contributes to the regulation of cargo passage through the secretory pathway.METHODS: The retention using selective hooks (RUSH) approach was used to image movement of model cargo (VSVg) between the ER and GA in various cell lines in which mTOR complexes were inhibited. We also investigated VSVg trafficking in TSC patient fibroblasts.
    RESULTS: We found that mTOR inhibition led to the overall enhancement of VSVg transport through the secretory pathway in PC12 cells and primary human fibroblasts. Also, in TSC1-deficient cells, VSVg transport was enhanced.
    CONCLUSIONS: Altogether, these data indicate the involvement of mTOR in the regulation of ER-to-GA cargo transport and suggest that impairments in exocytosis may be an additional cellular process that is disturbed in TSC.
    Keywords:  Endoplasmic reticulum; Golgi apparatus; MTOR; Retention using selective hooks; Tuberous sclerosis complex; VSVg
  4. Development. 2020 Jan 01. pii: dev.181727. [Epub ahead of print]
      In many eukaryotes, the small GTPase Rheb functions as a switch to toggle activity of TOR complex 1 (TORC1) between anabolism and catabolism, thus controlling lifespan, development, and autophagy. Our CRISPR-generated, fluorescently tagged endogenous C. elegans RHEB-1 and DAF-15/Raptor are expressed ubiquitously and localize to lysosomes. Disruption of LET-363/TOR and DAF-15/Raptor are required for development past the third larval stage (L3). We observed that deletion of RHEB-1 similarly conferred L3 arrest. Unexpectedly, robust RNAi-mediated depletion of TORC1 components caused arrest at stages prior to L3. Accordingly, conditional depletion of endogenous DAF-15/Raptor in the soma revealed that TORC1 is required at each stage of the life cycle to progress to the next stage. Reversal of DAF-15 depletion permits arrested animals to recover to continue development. Our results are consistent with TORC1 functioning as a developmental checkpoint that governs at each stage the decision of the animal to progress through development.
    Keywords:  Ral; RalGAP; TSC; Tuberous sclerosis complex; mTOR; mTORC1
  5. Childs Nerv Syst. 2021 May 18.
      INTRODUCTION: The association between cardiac rhabdomyoma and intraventricular tumors and/or subcortical nodules is characteristic of tuberous sclerosis complex (TSC). Patients with TSC may have refractory seizures, autistic behavior, and cognitive decline.CASE REPORT: The patient received the fetal diagnosis of TSC at the age of 19 weeks of gestations, where presented at prenatal ultrasound cardiac and brain tumors. Fetal MRI showed a lesion in the right and left lateral ventricles near the foramen of Monro associated with subependymal lesions along the entire ependyma of the lateral ventricles and several subcortical tubercles, and the fetal Doppler echocardiogram revealed three cardiac lesions. The fetus underwent intrauterine treatment with everolimus and presented regression and subsequent stabilization of the cardiac and brain lesions; additionally, the patient did not develop seizures or autism and presented good neuropsychomotor development.
    CONCLUSION: It is the first evidence that mTOR inhibitors may help to prevent neurological complications associated with TSC.
    Keywords:  Fetal brain tumor; Fetal chemotherapy; Tuberous sclerosis complex
  6. J Neurosurg Pediatr. 2021 May 21. pii: 2020.11.PEDS20823. [Epub ahead of print] 1-4
      Laser interstitial thermal therapy (LITT) is increasingly used as a surgical option for the treatment of epilepsy. Placement of the laser fibers relies on stereotactic navigation with cranial fixation pins. In addition, the laser fiber is stabilized in the cranium during the ablation using a cranial bolt that assumes maturity of the skull. Therefore, younger infants (< 2 years of age) have traditionally not been considered as candidates for LITT. However, LITT is an appealing option for patients with familial epilepsy syndromes, such as tuberous sclerosis complex (TSC), due to the multiplicity of lesions and the likely need for multiple procedures. A 4-month-old infant with TSC presented with refractory focal seizures despite receiving escalating doses of 5 antiepileptic medications. Electrographic and clinical seizures occurred numerous times daily. Noninvasive evaluations, including MRI, magnetoencephalography, scalp EEG, and SPECT, localized the ictal onset to a left frontal cortical tuber in the premotor area. In this paper, the authors report a novel technique to overcome the challenges of performing LITT in an infant with an immature skull by repurposing the Navigus biopsy skull mount for stereotactic placement of a laser fiber using electromagnetic-based navigation. The patient underwent successful ablation of the tuber and remained seizure free 4 months postoperatively. To the authors' knowledge, this is the youngest reported patient to undergo LITT. A safe method is described to perform LITT in an infant using commonly available tools without dedicated instrumentation beyond standard stereotactic navigation, a biopsy platform, and the Visualase system.
    Keywords:  LITT; MRI-guided laser interstitial thermal therapy; MRgLITT; ablation; epilepsy; oncology; surgical technique; tuber; tuberous sclerosis
  7. Front Pharmacol. 2021 ;12 647232
      The purposes of this study were to explore the population pharmacokinetics and initial dose optimization of sirolimus improving drug blood level for seizure control in pediatric patients with tuberous sclerosis complex (TSC). Eighty pediatric patients diagnosed with TSC-related epilepsy were included for analysis. Sirolimus concentrations, physiological and biochemical indexes, and drug combination were collected to build a nonlinear mixed effect (NONMEM) model. Initial dose optimization was simulated by the Monte Carlo method. The weight and concomitant medication of oxcarbazepine affected sirolimus clearance. Without oxcarbazepine, for once-daily sirolimus regimen, the doses of 0.07, 0.06, 0.05, 0.04, and 0.03 mg/kg/day were recommended for weights of 5-7.5, 7.5-11.5, 11.5-19, 19-40, and 40-70 kg, respectively; for twice-daily sirolimus regimen, the doses of 0.05, 0.04, and 0.03 were recommended for weights of 5-8, 8-20, and 20-70 kg, respectively. With oxcarbazepine, for once-daily sirolimus regimen, the doses of 0.09, 0.08, 0.07, 0.06, 0.05, and 0.04 mg/kg/day were recommended for weights of 5-7.5, 7.5-10, 10-13.5, 13.5-20, 20-35, and 35-70 kg, respectively; for twice-daily sirolimus regimen, the doses of 0.06, 0.05, 0.04, and 0.03 were recommended for weights of 5-7, 7-14.5, 14.5-38, and 38-70 kg, respectively. The present study was the first to establish a population pharmacokinetic model of sirolimus improving drug blood level for seizure control in pediatric patients with TSC and recommend the initial dosage regimen.
    Keywords:  initial dose optimization; population pharmacokinetics; seizure control; sirolimus; tuberous sclerosis complex
  8. Cell Commun Signal. 2021 May 20. 19(1): 57
      BACKGROUND: Cells adapt their metabolism and activities in response to signals from their surroundings, and this ability is essential for their survival in the face of perturbations. In tissues a deficit of these mechanisms is commonly associated with cellular aging and diseases, such as cardiovascular disease, cancer, immune system decline, and neurological pathologies. Several proteins have been identified as being able to respond directly to energy, nutrient, and growth factor levels and stress stimuli in order to mediate adaptations in the cell. In particular, mTOR, AMPK, and sirtuins are known to play an essential role in the management of metabolic stress and energy balance in mammals.METHODS: To understand the complex interactions of these signalling pathways and environmental signals, and how those interactions may impact lifespan and health-span, we have developed a computational model of metabolic signalling pathways. Specifically, the model includes (i) the insulin/IGF-1 pathway, which couples energy and nutrient abundance to the execution of cell growth and division, (ii) mTORC1 and the amino acid sensors such as sestrin, (iii) the Preiss-Handler and salvage pathways, which regulate the metabolism of NAD+ and the NAD+ -consuming factor SIRT1, (iv) the energy sensor AMPK, and (v) transcription factors FOXO and PGC-1α.
    RESULTS: The model simulates the interactions among key regulators such as AKT, mTORC1, AMPK, NAD+ , and SIRT, and predicts their dynamics. Key findings include the clinically important role of PRAS40 and diet in mTORC1 inhibition, and a potential link between SIRT1-activating compounds and premature autophagy. Moreover, the model captures the exquisite interactions of leucine, sestrin2, and arginine, and the resulting signal to the mTORC1 pathway. These results can be leveraged in the development of novel treatment of cancers and other diseases.
    CONCLUSIONS: This study presents a state-of-the-art computational model for investigating the interactions among signaling pathways and environmental stimuli in growth, ageing, metabolism, and diseases. The model can be used as an essential component to simulate gene manipulation, therapies (e.g., rapamycin and wortmannin), calorie restrictions, and chronic stress, and assess their functional implications on longevity and ageing-related diseases. Video Abstract.
    Keywords:  Ageing; Autophagy; Growth factor signaling; Longevity; MTOR; Metabolism; NAD+ ; Proliferation; Sirtuins; System biology
  9. J Cell Sci. 2020 Jan 01. pii: jcs.236661. [Epub ahead of print]
      Epithelial cells such as liver-resident hepatocytes rely heavily on the Rab family of small GTPases to perform membrane trafficking events that dictate cell physiology and metabolism. Not surprisingly, disruption of several Rabs can manifest in metabolic diseases or cancer. Rab32 is expressed in many secretory epithelial cells but its role in cellular metabolism is virtually unknown. In this study, we find that Rab32 associates with lysosomes and regulates proliferation and cell size of Hep3B hepatoma and HeLa cells. Specifically, we identify that Rab32 supports mTORC1 signaling under basal and amino acid stimulated conditions. Consistent with inhibited mTORC1, an increase in nuclear TFEB localization and lysosome biogenesis is also observed in Rab32-depleted cells. Finally, we find that Rab32 interacts with mTOR kinase and that loss of Rab32 reduces the association of mTOR and mTORC1 pathway proteins with lysosomes, suggesting that Rab32 regulates lysosomal mTOR trafficking. In summary, these findings suggest that Rab32 functions as a novel regulator of cellular metabolism through supporting mTORC1 signaling.
    Keywords:  Lysosome; MTORC1; S6K; Small Rab GTPase; TFEB
  10. J Cell Sci. 2020 Jan 01. pii: jcs.247817. [Epub ahead of print]
      In Schizosaccharomyces pombe, a general strategy for survival in response to environmental changes is sexual differentiation, which is triggered by TORC1 inactivation. However, mechanisms of TORC1 regulation in fission yeast remain poorly understood. In this study, we found that Pef1, which is an ortholog of mammalian CDK5, regulates the initiation of sexual differentiation through positive regulation of TORC1 activity. Conversely, deletion of pef1 leads to activation of autophagy and subsequent excessive TORC1 reactivation during the early phases of the nitrogen starvation response. This excessive TORC1 reactivation results in the silencing of the Ste11-Mei2 pathway and mating defects. Additionally, we found that pef1 genetically interacts with tsc1/2 in TORC1 regulation, and physically interacts with three types of cyclins, Clg1, Pas1, and Psl1. The double deletion of clg1 and pas1 promotes activation of autophagy and TORC1 during nitrogen starvation, similar to pef1Δ cells. Overall, our work suggests that Pef1-Clg1 and Pef1-Pas1 complexes regulate initiation of sexual differentiation through control of the TSC-TORC1 pathway and autophagy.
    Keywords:  Autophagy; CDK5; Cyclin; Pef1; Sexual differentiation; TORC1
  11. Adv Anat Pathol. 2021 Mar 29.
      Renal neoplasms largely favor male patients; however, there is a growing list of tumors that are more frequently diagnosed in females. These tumors include metanephric adenoma, mixed epithelial and stromal tumor, juxtaglomerular cell tumor, mucinous tubular and spindle cell carcinoma, Xp11.2 (TFE3) translocation-associated renal cell carcinoma, and tuberous sclerosis complex (somatic or germline) associated renal neoplasms. The latter category is a heterogenous group with entities still being delineated. Eosinophilic solid and cystic renal cell carcinoma is the best-described entity, whereas, eosinophilic vacuolated tumor is a proposed entity, and the remaining tumors are currently grouped together under the umbrella of tuberous sclerosis complex/mammalian target of rapamycin-related renal neoplasms. The entities described in this review are often diagnostic considerations when evaluating renal mass tissue on biopsy or resection. For example, Xp11.2 translocation renal cell carcinoma is in the differential when a tumor has clear cell cytology and papillary architecture and occurs in a young or middle-aged patient. In contrast, tuberous sclerosis complex-related neoplasms often enter the differential for tumors with eosinophilic cytology. This review provides an overview of the clinical, gross, microscopic, immunohistochemical, genetic, and molecular alterations in key renal neoplasms occurring more commonly in females; differential diagnoses are also discussed regardless of sex predilection.