A PTIP–PA1 subcomplex promotes transcription for IgH class switching independently from the associated MLL3/MLL4 methyltransferase complex

Abstract

Class switch recombination (CSR) diversifies antibodies for productive immune responses while maintaining stability of the B-cell genome. Transcription at the immunoglobulin heavy chain (Igh) locus targets CSR-associated DNA damage and is promoted by the BRCT domain-containing PTIP (Pax transactivation domain-interacting protein). Although PTIP is a unique component of the mixed-lineage leukemia 3 (MLL3)/MLL4 chromatin-modifying complex, the mechanisms for how PTIP promotes transcription remain unclear. Here we dissected the minimal structural requirements of PTIP and its different protein complexes using quantitative proteomics in primary lymphocytes. We found that PTIP functions in transcription and CSR separately from its association with the MLL3/MLL4 complex and from its localization to sites of DNA damage. We identified a tandem BRCT domain of PTIP that is sufficient for CSR and identified PA1 as its main functional protein partner. Collectively, we provide genetic and biochemical evidence that a PTIP-PA1 subcomplex functions independently from the MLL3/MLL4 complex to mediate transcription during CSR. These results further our understanding of how multifunctional chromatin-modifying complexes are organized by subcomplexes that harbor unique and distinct activities.

Replication fork stability confers chemoresistance in BRCA-deficient cells

“…To identify the region of PTIP that promotes RF degradation in Brca2 -deficient cells, we expressed EV (empty vector), FL (full-length PTIP), W165R (disrupting interactions with PA1) 25,26 , W663R (disrupting interactions with 53BP1 at DSBs) 25 or Del-BRCT5-6 (disrupting interaction with MLL3/4 independently of DSBs) 23,24,26 in Brca2/Ptip -doubly-deficient cells. We observed that only reconstitution of Brca2/Ptip -deficient cells with PTIP-Del-BRCT5-6 maintained fork protection (Extended Data Fig.…”

KMT2C/D COMPASS complex-associated diseases [KCDCOM-ADs]: an emerging class of congenital regulopathies

“…The K CD COMs' non-WRAD subunits are also more abundant than the KMT2 proteins and likewise are promiscuous in their protein interactions. Of note, the PTIP subunit is more abundantly found as a 53BP1complex member or as an independent heterodimer complexed with PA1 than as a member of K CD COM [32,42]. In addition, substoichiometric levels of NCOA6 and KDM6A protein were detected in HeLa cell K CD COM, suggesting that fewer than half of these complexes contained these specific subunits at steady-state.…”

BRCT-domain protein BRIT1 influences class switch recombination

“…The correlation between BRCT domains, per se, and molecular functions in CSR is not straightforward. For example, the N terminus BRCT domain of PTIP is required for CSR, whereas the C terminus BRCT repeats of 53BP1 are dispensable (37,38). BRIT1 contains an N-terminal BRCT domain and two C-terminal BRCT domains (Fig.…”