Probing chromatin function with NMR

The fascinating thing about chromatin is that fulfills two conflicting roles: on one hand it compacts the DNA, represses unwanted transcription, and protects our genetic information from damage, on the other hand it attracts a myriad of proteins to regulate DNA damage repair, replication, and gene transcription. How these proteins recognize, modify, assemble, remodel or disassemble nucleosomes, the fundamental building block of chromatin, is one of the key questions in chromatin biology.
Our lab focusses on solving the molecular mechanisms underlying chromatin function using an NMR-driven integrative structural biology approach. In this talk I will highlight our recent results in the study of nucleosome assembly during DNA repair. We uncovered that intrinsically disordered histone chaperone APLF can single-handedly assemble the histone octamer and transfer it to the DNA to form nucleosomes, in contrast to the prevailing model of stepwise assembly. I will further present our recent work on the mechanism of nucleosome remodeling by ATP-dependent chromatin remodeler ISWI. This protein machine can translocate the DNA in the nucleosome to alter the spacings between nucleosomes within chromatin. We show that the ISWI ATPase domain (75 kDa) is an intrinsically dynamic machine that causes wide-spread conformational changes and alteration of histone-DNA contacts in the nucleosomes. These data provide crucial support for the twist-diffusion mechanism of remodeling and highlight the nucleosome as a plastic, allosteric unit. Finally, I will present some of our experiences in solution NMR at 1.2 GHz.