Mapping ISG15 conjugation sites in THP1 cells during infection with Mycobacterium tuberculosis

ABSTRACT: ISG15 is an interferon-stimulated genes (ISGs) and an ubiquitin-like protein that undergoes conjugation to substrate proteins. Similar to ubiquitin conjugation, ISG15 conjugation is mediated by the ISG15 conjugation machinery. Recognition of Mycobacterium tuberculosis (Mtb) induces the accumulation of the ISG15 conjugation machinery on lipid droplets (LDs) in the cytosol of infected macrophages. This suggests that some ISG15 conjugation substrates might be recruited to the surface of lipid droplets. In this study, we identified the ISG15 conjugation substrates and modification sites in Mtb-infected macrophages. To do so, we enriched for ISG15- and ubiquitin-modified peptides with antibody raised against GlyGly(K) peptides from Mtb-infected THP1 cells. To distinguish ISG15- form ubiquitin-modification sites, we used ISG15-deficient macrophages with a constitutive ISG15 knockdown. GlyGly(K) modification sites found in control macrophages but absent in ISG15-lacking macrophages are likely bona fide ISG15-conjugation sites.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human) Mycobacterium Tuberculosis H37rv

TISSUE(S): Cell Culture

SUBMITTER: Fabien Thery  

LAB HEAD: Prof. Francis Impens

PROVIDER: PXD075338 | Pride | 2026-10-16

REPOSITORIES: Pride