Dataset Information

Basolateral Protein Scribble Provides Phosphatase PP1 to a Signaling Network Maintaining Apicobasal Polarity

ABSTRACT: Scribble, a member of the LAP protein family, contributes to the apicobasal polarity (ABP) of epithelial cells. The LAP unique region (LUR) of these proteins is essential for ABP function, and includes a highly conserved Leucine Reach Repeat (LRR) domain. Here we study how the LRR domain of Scribble maintains ABP. We show that its concave surface participates in three types of mutually exclusive interactions 1. homodimerization serving as an auto-inhibitory mechanism, 2. interactions with a diverse set of polarity proteins, such as Llgl1, Llgl2, EPB41L2 and EPB41L5, which produce distinct multiprotein complexes, and 3. a direct interaction with the protein phosphatase, PP1, which forms a dimeric LRR domain-PP1 complex. Our results suggest that the latter complex maintains PP1 in the basolateral cell cortex in close proximity to PP1 targets. Such organization generates a dynamic network where PP1 could be immediately dispatched from the Scribble-PP1 complex to particular protein ligands. This mechanism for controlling dephosphorylation kinetics of phosphorylated proteins could be universal for all members of the LAP protein family, which includes Erbin, Lano, and the neuron-specific protein, Densin-180.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Colon

DISEASE(S): Colon Cancer

SUBMITTER: Indrajyoti Indra

LAB HEAD: Sergey Troyanovsky

PROVIDER: PXD028919 | Pride | 2021-11-03

REPOSITORIES: Pride

ACCESS DATA

Publications

Basolateral protein Scribble binds phosphatase PP1 to establish a signaling network maintaining apicobasal polarity.

Troyanovsky Regina B RB Indra Indrajyoti I Kato Rei R Mitchell Brian J BJ Troyanovsky Sergey M SM

The Journal of biological chemistry 20211008 5

Scribble, a member of the LAP protein family, contributes to the apicobasal polarity (ABP) of epithelial cells. The LAP-unique region of these proteins, which is essential and sufficient for ABP, includes a conserved Leucine-Rich Repeat (LRR) domain. The major binding partners of this region that could regulate ABP remain unknown. Here, using proteomics, native gel electrophoresis, and site-directed mutagenesis, we show that the concave surface of LRR domain in Scribble participates in three typ ...[more]

PMID: 34634305

Similar Datasets

Project description:BACKGROUND & AIMS- More frequent interaction of bacteria with the colonic epithelium is associated with ulcerative colitis (UC). The identities of all proteins which promote bacterial clearance in colonic epithelial cells are unknown. Previously, we discovered that dCAP-D3 (Chromosome Associated Protein-D3), regulates responses to bacterial infection. We examined whether CAP-D3 promotes bacterial clearance in human colonic epithelium. METHODS- Clearance of Salmonella or adherent-invasive Escherichia coli LF82 was assessed by gentamycin protection assays in HT-29 and Caco-2 cells expressing CAP-D3 shRNA. CAP-D3 levels in colonic epithelial cells from healthy and UC patient tissues were analyzed by immunoblot. RNA-sequencing identified bacterially-induced CAP-D3 target genes. The role of CAP-D3 target genes in bacterial clearance was analyzed by gentamycin protection assays, immunofluorescent staining, and by using pharmacologic inhibitors. RESULTS- CAP-D3 expression was reduced in colonic epithelial cells from UC patients with active disease. Reduction of CAP-D3 expression inhibited autophagy and decreased intracellular bacterial clearance. The components of the heterodimeric SLC7A5/SLC3A2 amino acid transporter were identified as CAP-D3 target genes; their levels increased in infected, CAP-D3 deficient cell lines and in cells from UC patients. In HT-29 cells, this resulted in earlier SLC7A5 recruitment to Salmonella-containing vacuoles, increased mTOR activity, and enhanced bacterial survival. Inhibition of SLC7A5/SLC3A2 or mTOR activity rescued the bacterial clearance defect in CAP-D3 deficient cells. CONCLUSIONS- CAP-D3 attenuates amino acid transporter transcription to promote bacterial autophagy in colon epithelial cells. CAP-D3 protein levels are decreased in patients with active UC, suggesting that CAP-D3 is a potential therapeutic target to restore mucosal homeostasis in UC patients. Three RNA samples from 3 independent experiments including timepoints taken at 0, 0.5 and 7 hours post-infection were analyzed on a bioanalyzer for quality; one of the 0.5 hour post-infection samples was excluded at this time due to poor RNA purity. Directional, cDNA libraries made from cellular mRNAs were generated from the other 8 samples and sequenced (paired-end sequencing of 100 bp reads) in the Genomics Core at the University of Chicago on an Illumina HiSeq2000.

Dataset Information

Basolateral Protein Scribble Provides Phosphatase PP1 to a Signaling Network Maintaining Apicobasal Polarity

Publications

Basolateral protein Scribble binds phosphatase PP1 to establish a signaling network maintaining apicobasal polarity.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure