Research Summary
Our goals are to delineate the mechanisms of DNA repair processes
in eukaryotes, and to determine how damage bypass occurs during DNA
replication. We have identified key proteins responsible for
post-replicative bypass of damaged DNA. One is Rad6-Rad18, in which an
ubiquitin-conjugating enzyme (Rad6) exists in tight association with a
DNA binding protein (Rad18), providing a mechanism for the targeting of
ubiquitin-conjugating activity to damage sites in DNA. DNA polymerase η
, which has the unusual ability to replicate through cyclobutane
pyrimidine dimers (CPDs), was identified in our group; we showed that
its mutational inactivation is the cause of the cancer-prone syndrome,
the variant form of xeroderma pigmentosum in humans. Interestingly, Pol?
replicates through a CPD with the same accuracy and efficiency as it
replicates through undamaged DNAs.
Biochemical studies carried out in our group and structural
studies done in collaboration with Aneel Aggarwal at Mt. Sinai School of
Medicine, NY have revealed a high degree of specificity in the manner
by which the various yeast and human translesion synthesis DNA
polymerases accomplish lesion bypass. For example, Polη , which is
specialized for replicating through CPDs, has the unique ability to hold
two templating residues in its active site. DNA polymerases κ and ζ,
on the other hand, are uniquely adapted for extending mispaired primer
termini and for carrying out the extension step of lesion bypass, in
which they extend from the nucleotide inserted opposite the lesion site
by a DNA polymerase such as ι or Rev1. Pols κ and ζ differ in the types
of lesions from which they can extend in very distinct and remarkable
ways, and biochemical studies are revealing some highly unusual features
that these polymerases adopt in performing their tasks.
Our combined genetic, biochemical, and structural approaches have
yielded insights into the novel mechanisms of DNA synthesis adopted by
the various yeast and human translesion synthesis DNA polymerases. We
are now determining the in vivo contributions of these polymerases to
error-free vs. mutagenic lesion bypass, and we are examining the
mechanisms by which translesion synthesis DNA polymerases gain access to
the replication fork and mediate lesion bypass.
Selected Publications
- Silverstein, T. D., R. E. Johnson, R. Jain, L. Prakash, S. Prakash, and A. K. Aggarwal (2010) Structural basis for the suppression of skin cancers by DNA polymerase eta. Nature 465: 1039-1043.
- Yoon, J.-H., G. Bhatia, S. Prakash, and L. Prakash (2010) Error-free replicative bypass of thymine glycol by the combined action of DNA polymerases k and z in human cells. Proc. Natl. Acad. Sci. 107:14116-14122.
- Gangavarapu V, Santa Maria SR, Prakash S, Prakash L. Requirement of replication checkpoint protein kinases mec1/rad53 for postreplication repair in yeast. Mbio. 2011: 2(3): e00079-11
- Ummat A, Rechkoblit O, Jain R, Roy Choudhury J, Johnson RE, Silverstein TD, Buku A, Lone S, Prakash L, Prakash S, Aggarwal AK. Structural basis for cisplatin DNA damage tolerance by human polymerase h during cancer chemotherapy. Nature Struct. Mol. Biol. 2012:19(6):628-632.
- Yoon J-H, Prakash S, Prakash L. Requirement of Rad18 protein for replication through DNA lesions in mouse and human cells. Proc. Natl. Acad. Sci. 2012:109(20):7799-7804.
- Yoon J-H, Prakash S, Prakash L. Genetic control of translesion synthesis on leading and lagging DNA strands in plasmids derived from Epstein-Barr virus in human cells. mBio. 2012: 3(5):e00271-12.
- Gómez-Llorente Y, Malik R, Jain R, Roy Choudhury J, Johnson RE, Prakash L, Prakash S, Ibarretxena-Belandia I, Aggarwal AK. The architecture of yeast DNA polymerase z. Cell Reports 2013:5:79-86.
- Yoon J-H, Roy Choudhury J, Park J, Prakash S, Prakash L. A role for DNA polymerase q in promoting replication through oxidative DNA lesion, thymine glycol, in human cells. J. Biol. Chem. 2014:289:13177-13185.
- Yoon J-H, Acharya N, Park J, Basu D, Prakash S, Prakash L. Identification of two functional PCNA-binding domains in human DNA polymerase k. Genes to Cell 2014:19:594-601.
- Rechkoblit, O., Gupta, Y. K., Malik, R., Rajashankar, R., Johnson, R. E., Prakash, L., Prakash, S., and Aggarwal, A. K. Structure and mechanism of human PrimPol, a DNA polymerase with primase activity. Science Advances 2016: 2:e1601317