Cellular mechanisms of DNA replication initiation

Life depends on the faithful transmission of genetic information from parent to progeny. Cellular life stores genetic content in the form of DNA and cells contain machinery that copies and disseminates duplicated DNA to daughter cells during cell division. Although the molecules which ready DNA for replication are well known, how their action is coordinated in cellular space and time remain a mystery. We are using live-cell fluorescence microscopy to directly visualize the early steps of DNA replication in human and fruity fly cells. The information we glean from cellular studies directly guides our efforts to develop an in vitro reconstituted system as a means to further dissect the molecular underpinnings of this fundamental process.

Speciailization of initiator disordered domains in protein phase separation

The metazoan DNA replication initiation factors (ORC, Cdc6 and Cdt1) contain extended regions of intrinsic disorder that facilitate chromatin association in cells and drive DNA-dependent self-assembly and phase separation in vitro. Biomolecular condensates that form upon initiator-DNA binding selectively recruit replication factors but exclude non-partner phase separating proteins. Interestingly, initiator-type disordered domains form a sequence class all their own. We seek to understand how this set of sequences facilitates DNA-dependent phase separation and aim to uncover the molecular basis for the inherent sorting capacity of initiator condensates.

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The molecular language of inter-IDR communication

There are thousands of extended (> 100 residues) intrinsically disordered regions across a given metazoan proteome. It is becoming increasingly clear that many biomolecular condensates rely on disordered sequences for the multivalent interactions that drive their assembly and for selective inter-IDR interactions that tune condensate composition. However, the language(s) of inter-IDR communication is largely a mystery. We are investigating the molecular determinants of inter-IDR interactions in an unbiased and proteome-wide fashion with the ultimate goal of developing algorithms for a priori prediction of inter-IDR interaction and  condensate sorting.

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