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Association of crumbs homolog-2 with mTORC1 in developing podocyte
Sho Hamano1, Yukino Nishibori1, Ichiro Hada1
1Department of Pediatrics, Kyorin University School of Medicine, Mitaka, Tokyo, Japan.
Abstract:
The evidence that gene mutations in the polarity determinant Crumbs homologs-2 (CRB2) cause congenital nephrotic syndrome suggests the functional importance of this gene product in podocyte development. Because another isoform, CRB3, was reported to repress the mechanistic/mammalian target of the rapamycin complex 1 (mTORC1) pathway, we examined the role of CRB2 function in developing podocytes in relation to mTORC1. In HEK-293 and MDCK cells constitutively expressing CRB2, we found that the protein localized to the apicolateral side of the cell plasma membrane and that this plasma membrane assembly required N-glycosylation. Confocal microscopy of the neonate mouse kidney revealed that both the tyrosine-phosphorylated form and non-phosphorylated form of CRB2 commence at the S-shaped body stage at the apicolateral side of podocyte precursor cells and move to foot processes in a capillary tuft pattern. The pattern of phosphorylated mTOR in developing podocytes was similar to that of CRB2 tyrosine phosphorylation. Additionally, the lack of a tyrosine phosphorylation site on CRB2 led to the reduced sensitivity of mTORC1 activation in response to energy starvation. CRB2 may play an important role in the mechanistic pathway of developing podocytes through tyrosine phosphorylation by associating with mTORC1 activation.
Insights
Crumbs homolog 2 (CRB2) protein is crucial for developing kidney podocytes. CRB2 associates with mTORC1 activation, impacting kidney development and potentially congenital nephrotic syndrome.
Area of Science:
- Cell Biology
- Developmental Biology
- Nephrology
Background:
- Mutations in Crumbs homolog 2 (CRB2) are linked to congenital nephrotic syndrome, highlighting its importance in podocyte development.
- CRB3, an isoform of CRB2, is known to inhibit the mechanistic/mammalian target of the rapamycin complex 1 (mTORC1) pathway.
Purpose of the Study:
- To investigate the role of CRB2 in developing podocytes, specifically its relationship with the mTORC1 pathway.
- To determine the localization and post-translational modifications of CRB2 during kidney development.
Main Methods:
- Expression of CRB2 in HEK-293 and MDCK cells to study protein localization and N-glycosylation requirements.
- Confocal microscopy of neonatal mouse kidneys to track CRB2 and phosphorylated mTOR patterns during podocyte development.
- Analysis of CRB2 tyrosine phosphorylation site mutants to assess mTORC1 activation sensitivity.
Main Results:
- CRB2 localizes to the apicolateral plasma membrane in a process dependent on N-glycosylation.
- CRB2 and phosphorylated mTOR exhibit similar spatiotemporal patterns in developing podocytes.
- Absence of a CRB2 tyrosine phosphorylation site reduces mTORC1 activation sensitivity during energy starvation.
Conclusions:
- CRB2 plays a significant role in developing podocytes.
- CRB2's tyrosine phosphorylation is linked to mTORC1 pathway activation, suggesting a mechanism for its function in kidney development.
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