Phosphoproteomic profiling highlights CDC42 and CDK2 as key players in the regulation of the TGF-β pathway in ALMS1 and BBS1 knockout models.

This research studied cells in the laboratory to better understand how Bardet-Biedl Syndrome (BBS) affects kidney function. The scientists used a gene-editing tool called CRISPR to remove the BBS1 gene from cells, then studied how this change affected important cell processes. They focused on a biological pathway called TGF-β, which is known to play a major role in the kidney disease that often develops in people with BBS and related conditions. The researchers found that when the BBS1 gene was missing, it disrupted the TGF-β pathway and affected how proteins in the cells were activated. They identified a protein called CDK2 as particularly important in BBS cells, and another protein called CDC42 in a related condition called Alström syndrome. These proteins appear to be key players in the chain reaction of cellular problems that may eventually lead to kidney disease in patients. This is early-stage laboratory research using cells rather than testing treatments in patients. However, understanding these specific protein pathways could be important for the future because it gives scientists potential targets for developing treatments. If researchers can find ways to correct the problems with CDK2 and the TGF-β pathway, it might help prevent or slow down the kidney disease that affects many people with BBS as they get older.

The primary cilium is a sensory organelle that extends from the plasma membrane. It plays a vital role in physiological and developmental processes by controlling different signalling pathways such as WNT, Sonic hedgehog (SHh), and transforming growth factor β (TGF-β). Ciliary dysfunction has been related to different pathologies such as Alström (ALMS) or Bardet-Biedl (BBS) syndrome. The leading cause of death in adults with these syndromes is chronic kidney disease (CKD), which is characterised by fibrotic and inflammatory processes often involving the TGF-β pathway. Using genomic editing with CRISPR-CAS9 and phosphoproteomics we have studied the TGF-β signalling pathway in knockout (KO) models for ALMS1 and BBS1 genes. We have developed a network diffusion-based analysis pipeline to expand the data initially obtained and to be able to determine which processes were deregulated in TGF-β pathway. Finally, we have analysed protein-protein and kinase-substrate interactions to prioritise candidate genes in the regulation of the TGF-β pathway in ALMS and BBS. Analysis of differentially phosphorylated proteins identified 10 candidate proteins in the ALMS1 KO model and 41 in the BBS1 KO model. After network expansion using a random walk with a restart algorithm, we were able to identify the TGF-β signalling pathway together with other related processes such as endocytosis in the case of ALMS1 or the regulation of the extracellular matrix in BBS1. Protein interaction analyses demonstrated the involvement of CDC42 as a central protein in the interactome in ALMS1 and CDK2 in the case of BBS1. In conclusion, the depletion of ALMS1 and BBS1 affects the TGF-β signalling pathway, conditioning the phosphorylation and activation of several proteins, including CDC42 in the case of ALMS1 and CDK2 in the case of BBS1.