Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2020 Nov 26;8:599890. doi: 10.3389/fcell.2020.599890

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

Copyright © 2020 Li and Niswander.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

PMC Copyright notice

TMEM132A stabilizes Wnt ligands and increases their binding with WLS. (A) Loss of TMEM132A destabilizes WNT protein levels. (Left panel) Scramble or TMEM132A siRNAs were transfected into HEK 293T cells followed by transfection with FL WNT3A or WNT5A-HA. After 24 hours the cells were treated with 100 μg/ml CHX and the protein levels of WNT3A or WNR5A were evaluated at the indicted time periods by Western blot. (Middle panel) TMEM132A knockdown efficiency before CHX treatment (t = 0). GAPDH served as an internal control. (Right panel) Relative WNT3A (upper panel) or WNT5A protein (lower panel) remained after CHX treatment was calculated by measuring the Western blot band grayscale. (B) WNT glycosylation is not altered by TMEM132A depletion. HEK 293T cells were co-transfected with TMEM132A siRNAs and WNT3A or WNT5A-HA, and the glycosylation status of WNT3A or WNT5A was determined by Western blot. Tun at the indicated concentrations was used to block global glycosylation as a positive control. Relative band intensity is given below corresponding panel (normalized to GADPH). (C) WNT palmitoylation is not altered by TMEM132A depletion. HEK 293T cells co-transfected with scramble or TMEM132A specific siRNAs and FL WNT3A or WNT5A-HA were labeled with 100 μM palmitic acid alkyne (Alk-C16:0), lysed and subjected to IP and on-bead click chemistry reaction in the presence of biotin azide plus to determine WNT3A/5A palmitoylation level by Western blot. TMEM132A knockdown efficiency is shown, with GAPDH as an internal control. Relative band intensity is given below corresponding panel (normalized to GADPH except the α-Biotin panel which is normalized to α-HA signal). (D) Co-IP was performed to determine the interaction between WLS and WNT3A or WNT5A in the presence or absence of overexpressed TMEM132A in HEK 293T cells. The result shows enhanced WLS-WNT3A or WNT5A interaction by addition of TMEM132A. Relative band intensity is given below corresponding panel (normalized to α-Myc signal) (E) Co-IP of FL WNT3A-HA and WNT5A-HA by FL T132A-Flag in HEK 293T cells.