About

William K. Holloman, Ph.D., is a Professor of Microbiology and Immunology at Weill Cornell Medicine. His research focuses on maintaining genomic stability through homologous recombination and DNA repair. His work investigates the molecular mechanisms of DNA repair processes, particularly the role of Rad52 in DNA repair, and how proteins involved in these processes are conserved across evolution from microbes to humans. Holloman's research explores the paradoxical functions of Rad52 in higher organisms, especially its potential role in overcoming barriers to DNA replication, and aims to understand the regulation of key proteins such as Rad51, BRCA2, and Dss1. His laboratory employs the yeast Ustilago maydis as a model organism to study these components, with the goal of gaining insights into how the human BRCA2 protein operates as a tumor suppressor to maintain genomic stability.

Research topics

• Biology
• Genetics
• Cell biology

Selected publications

• An antimorphic mutant of Brh2 resulting from a conservative amino acid change within the BRC element

  DNA repair · 2026-04-30

  articleSenior author
  PublisherDOI

• Dss1 facilitates Rad51 recruitment downstream of BRCA2/Brh2 in response to DNA damage

  DNA repair · 2026-01-23

  articleOpen access

  Homologous recombination (HR) is a major pathway for repair of DNA double-strand breaks (DSB), recovery of broken replication forks and formation of meiotic crossovers. HR provides a mechanism to precisely repair damaged DNA in a template-dependent process. The defining step in HR is homologous strand exchange directed by the RecA-related recombinase Rad51. BRCA2 and Brh2, the BRCA2 orthologue in Ustilago maydis, enable recombinational repair of DNA by controlling Rad51. In turn, Dss1, a small intrinsically disordered protein that binds to the C-terminal region of BRCA2/Brh2, regulates BRCA2/Brh2. In the present study, we dissect the interdependency of HR proteins for recruitment to DNA-damage induced foci using fluorescence microscopy and genetics. In U. maydis, Brh2 and Dss1 colocalize at DNA damage-induced foci. Dss1 recruitment to foci is dependent on interaction with full-length Brh2 and Dss1-Brh2 interaction is required for resistance to DNA damage. Further, Dss1 is required for Rad51 and Rec2 focus formation. Interestingly, we find that Rad52 is required for Brh2, Rec2 and Dss1 focus formation. In avian DT40 cells, we likewise show that endogenously tagged DSS1 redistributes into subnuclear foci in response to DNA damaging agents. However, DSS1 foci rarely colocalize with BRCA2 foci. Finally, Dss1 focus formation is inhibited by treatment with the proteasome inhibitor MG132, in both U. maydis and DT40 cells, suggesting a role of ubiquitin in homology-dependent repair.

  PublisherDOI

• DNA polymerase ζ has robust reverse transcriptase activity relative to other cellular DNA polymerases

  Journal of Biological Chemistry · 2024-10-23 · 7 citations

  articleOpen access

  Cell biology and genetic studies have demonstrated that DNA double-strand break (DSB) repair can be performed using an RNA transcript that spans the site of the DNA break as a template for repair. This type of DSB repair requires a reverse transcriptase to convert an RNA sequence into DNA to facilitate repair of the break, rather than copying from a DNA template as in canonical DSB repair. Translesion synthesis (TLS) DNA polymerases (Pol) are often more promiscuous than DNA Pols, raising the notion that reverse transcription could be performed by a TLS Pol. Indeed, several studies have demonstrated that human Pol η has reverse transcriptase activity, while others have suggested that the yeast TLS Pol ζ is involved. Here, we purify all seven known nuclear DNA Pols of Saccharomyces cerevisiae and compare their reverse transcriptase activities. The comparison shows that Pol ζ far surpasses Pol η and all other DNA Pols in reverse transcriptase activity. We find that Pol ζ reverse transcriptase activity is not affected by RPA or RFC/PCNA and acts distributively to make DNA complementary to an RNA template strand. Consistent with prior S. cerevisiae studies performed in vivo, we propose that Pol ζ is the major DNA Pol that functions in the RNA-templated DSB repair pathway.

  PublisherOA PDFDOI

• DNA polymerase ζ is a robust reverse transcriptase

  bioRxiv (Cold Spring Harbor Laboratory) · 2024-09-28

  preprintOpen accessCorresponding

  Abstract Cell biology and genetic studies have demonstrated that DNA double strand break (DSB) repair can be performed using an RNA transcript that spans the site of the DNA break as a template for repair. This type of DSB repair requires a reverse transcriptase to convert an RNA sequence into DNA to facilitate repair of the break, rather than copying from a DNA template as in canonical DSB repair. Translesion synthesis (TLS) DNA polymerases (Pol) are often more promiscuous than DNA Pols, raising the notion that reverse transcription could be performed by a TLS Pol. Indeed, several studies have demonstrated that human Pol η has reverse transcriptase activity, while others have suggested that the yeast TLS Pol ζ is involved. Here, we purify all seven known nuclear DNA Pols of Saccharomyces cerevisiae and compare their reverse transcriptase activities. The comparison shows that Pol ζ far surpasses Pol η and all other DNA Pols in reverse transcriptase activity. We find that Pol ζ reverse transcriptase activity is not affected by RPA or RFC/PCNA and acts distributively to make DNA complementary to an RNA template strand. Consistent with prior S. cerevisiae studies performed in vivo , we propose that Pol ζ is the major DNA Pol that functions in the RNA templated DSB repair pathway.

  PublisherDOI

• Ustilago maydis Trf2 ensures genome stability by antagonizing Blm-mediated telomere recombination: Fine-tuning DNA repair factor activity at telomeres through opposing regulations

  PLoS Genetics · 2024-12-09 · 1 citations

  articleOpen accessCorresponding

  TRF2 is an essential and conserved double-strand telomere binding protein that stabilizes chromosome ends by suppressing DNA damage response and aberrant DNA repair. Herein we investigated the mechanisms and functions of the Trf2 ortholog in the basidiomycete fungus Ustilago maydis, which manifests strong resemblances to metazoans with regards to the telomere and DNA repair machinery. We showed that UmTrf2 binds to Blm in vitro and inhibits Blm-mediated unwinding of telomeric DNA substrates. Consistent with a similar inhibitory activity in vivo, over-expression of Trf2 induces telomere shortening, just like deletion of blm, which is required for efficient telomere replication. While the loss of Trf2 engenders growth arrest and multiple telomere aberrations, these defects are fully suppressed by the concurrent deletion of blm or mre11 (but not other DNA repair factors). Over-expression of Blm alone triggers aberrant telomere recombination and the accumulation of aberrant telomere structures, which are blocked by concurrent Trf2 over-expression. Together, these findings highlight the suppression of Blm as a key protective mechanism of Trf2. Notably, U. maydis harbors another double-strand telomere-binding protein (Tay1), which promotes Blm activity to ensure efficient replication. We found that deletion of tay1 partially suppresses the telomere aberration of Trf2-depleted cells. Our results thus point to opposing regulation of Blm helicase by telomere proteins as a strategy for optimizing both telomere maintenance and protection. We also show that aberrant transcription of both telomere G- and C-strand is a recurrent phenotype of telomere mutants, underscoring another potential similarity between double strand breaks and de-protected telomeres.

  PublisherOA PDFDOI

• <i>Ustilago maydis</i> Trf2 ensures genome stability by antagonizing Blm-mediated telomere recombination: fine-tuning DNA repair factor activity at telomeres through opposing regulations

  bioRxiv (Cold Spring Harbor Laboratory) · 2023-04-17

  preprintOpen access

  Abstract TRF2 is an essential and conserved double-strand telomere binding protein that stabilizes chromosome ends by suppressing DNA damage response and aberrant DNA repair. Herein we investigated the mechanisms and functions of the Trf2 ortholog in the basidiomycete fungus Ustilago maydis, which manifests strong resemblances to metazoans with regards to the telomere and DNA repair machinery. We showed that Um Trf2 binds to Blm in vitro and inhibits Blm-mediated unwinding of telomeric DNA substrates. Consistent with a similar inhibitory activity in vivo , over-expression of Trf2 induces telomere shortening, just like deletion of blm, which is required for efficient telomere replication. While the loss of Trf2 engenders growth arrest and multiple telomere aberrations, these defects are fully suppressed by the concurrent deletion of blm or mre11 (but not other DNA repair factors). Over-expression of Blm alone triggers aberrant telomere recombination and the accumulation of aberrant telomere structures, which are blocked by concurrent Trf2 over-expression. Together, these findings highlight the suppression of Blm as a key protective mechanism of Trf2. Notably, U. maydis harbors another double-strand telomere-binding protein (Tay1), which promotes Blm activity to ensure efficient replication. We found that deletion of tay1 partially suppresses the telomere aberration of Trf2-depleted cells. Our results thus point to opposing regulation of Blm helicase by telomere proteins as a strategy for optimizing both telomere maintenance and protection. We also show that aberrant transcription of both telomere G- and C-strand is a recurrent phenotype of telomere mutants, underscoring another potential similarity between double strand breaks and de-protected telomeres. Author Summary The ends of linear chromosomes are protected from abnormal repair by a collection of telomere proteins. One protein that plays an especially prominent role is TRF2, which binds to double-stranded telomere repeats. In this study, we analyzed the mechanisms and functions of Trf2 in a yeast-like fungus named Ustilago maydis , which manifests a high degree of similarity to animal cells with respect to telomere regulation. We showed that Trf2 binds directly to a conserved DNA helicase called Blm and inhibits the ability of Blm to unwind telomeric DNA in a purified, cell-free reaction. We also used over-expression and depletion of either Trf2 or Blm or both to demonstrate an inhibitory effect of Trf2 on Blm function in vivo . For example, depletion of Trf2 triggers Blm-dependent telomere aberrations and cell death. Interestingly, another double-strand telomere binding protein named Tay1 was found to stimulate Blm activity to promote telomere replication. Together, our results indicate that U. maydis optimizes Blm function through opposing regulation of its activity via distinct telomere proteins. We also detected high levels of abnormal transcripts that correspond to both strands of telomeres in a variety of telomere mutants, suggesting that de-protected telomeres are permissive substrates for the transcription apparatus.

  PublisherOA PDFDOI

• Determinants governing BRC function evaluated by mutational analysis of Brh2 in Ustilago maydis

  DNA repair · 2023-04-26 · 8 citations

  articleSenior author
  PublisherDOI

• Ustilago maydis telomere protein Pot1 harbors an extra N-terminal OB fold and regulates homology-directed DNA repair factors in a dichotomous and context-dependent manner

  PLoS Genetics · 2022-05-19 · 8 citations

  articleOpen access

  The telomere G-strand binding protein Pot1 plays multifaceted roles in telomere maintenance and protection. We examined the structure and activities of Pot1 in Ustilago maydis, a fungal model that recapitulates key features of mammalian telomere regulation. Compared to the well-characterized primate and fission yeast Pot1 orthologs, UmPot1 harbors an extra N-terminal OB-fold domain (OB-N), which was recently shown to be present in most metazoans. UmPot1 binds directly to Rad51 and regulates the latter's strand exchange activity. Deleting the OB-N domain, which is implicated in Rad51-binding, caused telomere shortening, suggesting that Pot1-Rad51 interaction facilitates telomere maintenance. Depleting Pot1 through transcriptional repression triggered growth arrest as well as rampant recombination, leading to multiple telomere aberrations. In addition, telomere repeat RNAs transcribed from both the G- and C-strand were dramatically up-regulated, and this was accompanied by elevated levels of telomere RNA-DNA hybrids. Telomere abnormalities of pot1-deficient cells were suppressed, and cell viability was restored by the deletion of genes encoding Rad51 or Brh2 (the BRCA2 ortholog), indicating that homology-directed repair (HDR) proteins are key mediators of telomere aberrations and cellular toxicity. Together, these observations underscore the complex physical and functional interactions between Pot1 and DNA repair factors, leading to context-dependent and dichotomous effects of HDR proteins on telomere maintenance and protection.

  PublisherDOI

• <i>Ustilago maydis</i> telomere protein Pot1 harbors an extra N-terminal OB fold and regulates homology-directed DNA repair factors in a dichotomous and context-dependent manner

  bioRxiv (Cold Spring Harbor Laboratory) · 2021-09-07

  preprintOpen access

  Abstract The telomere G-strand binding protein Pot1 plays multifaceted roles in telomere maintenance and protection. We examined the structure and activities of Pot1 in Ustilago maydis , a fungal model that recapitulates key features of mammalian telomere regulation. Compared to the well-characterized primate and fission yeast Pot1 orthologs, Um Pot1 harbors an extra N-terminal OB-fold domain (OB-N), which was recently shown to be present in most metazoans. Um Pot1 binds directly to Rad51 and regulates the latter’s strand exchange activity. Deleting the OB-N domain, which is implicated in Rad51-binding, caused telomere shortening, suggesting that Pot1-Rad51 interaction facilitates telomere replication. Depleting Pot1 through transcriptional repression triggered growth arrest as well as rampant recombination, leading to multiple telomere aberrations. In addition, telomere repeat RNAs transcribed from both the G- and C-strand were dramatically up-regulated, and this was accompanied by elevated levels of telomere RNA-DNA hybrids. Telomere abnormalities of pot1 -deficient cells were suppressed, and cell viability was restored by the deletion of genes encoding Rad51 or Brh2 (the BRCA2 ortholog), indicating that homology-directed repair (HDR) proteins are key mediators of telomere aberrations and cellular toxicity. Together, these observations underscore the complex physical and functional interactions between Pot1 and DNA repair factors, leading to context-dependent and dichotomous effects of HDR proteins on telomere maintenance and protection.

  PublisherOA PDFDOI

• Structurally distinct duplex telomere repeat-binding proteins in <i>Ustilago maydis</i> execute specialized, non-overlapping functions in telomere recombination and telomere protection

  bioRxiv (Cold Spring Harbor Laboratory) · 2020-11-07 · 1 citations

  preprintOpen access

  Abstract Duplex telomere binding proteins exhibit considerable structural and functional diversity in different phyla. Herein we address the distinct properties and functions of two Myb-containing, duplex telomere repeat-binding factors in Ustilago maydis , a basidiomycete fungus that is evolutionarily distant from the standard budding and fission yeasts. The two telomere-binding proteins in U. maydis , named Um Trf1 and Um Trf2, have different domain organizations and belong to distinct protein families with different phylogenetic distributions. Despite these differences, they exhibit comparable affinities and similar sequence specificity for the canonical, 6-base-pair telomere repeats. Deletion of trf1 triggers preferential loss of long telomere tracts, suggesting a role for the encoded protein in promoting telomere replication. Trf1 loss also partially suppresses the ALT-like phenotypes of ku70 -deficient mutants, suggesting a novel role for a telomere protein in stimulating ALT-related pathways. In keeping with these ideas, we found that purified Trf1 can modulate the helicase activity of Blm, a conserved telomere replication and recombination factor. In contrast, trf2 appears to be essential and transcriptional repression of this gene leads to severe growth defects and profound telomere aberrations that encompass telomere length heterogeneity, accumulation of extrachromosomal telomere repeats such as C-circles, and high levels of single-stranded telomere DNA. These observations support a critical role for Um Trf2 in telomere protection. Together, our findings point to a unique, unprecedented division of labor between the two major duplex telomere repeat-binding factors in Ustilago maydis . Comparative analysis of Um Trf1 homologs in different phyla reveals a high degree of functional diversity for this protein family, and provides a case study for how a sequence-specific DNA binding protein can acquire and lose functions at different chromosomal locations.

  PublisherOA PDFDOI

Recent grants

• NIH Grant R01GM036327

  NIH · $517k · 1988

• NIH Grant R01GM042548

  NIH · $346k · 1993

• NIH Grant R01GM027103

  NIH · $53k · 1985

• NIH Grant R01GM053732

  NIH · $655k · 1999

• NIH Grant R01GM042482

  NIH · $7.2M · 2015

Frequent coauthors

• Jeanette Sutherland

  Cornell University

  77 shared

• Neal F. Lue

  Weill Cornell Medicine

  62 shared

• Syed Zahid

  Hearst (United States)

  40 shared

• Sarah Aloe

  Cornell University

  36 shared

• Milorad Kojic

  University of Belgrade

  33 shared

• Eric B. Kmiec

  Christiana Care Health System

  31 shared

• Eun Young Yu

  Hearst (United States)

  24 shared

• Qingwen Zhou

  Cornell University

  23 shared

Education

• Ph.D, Biochemistry

  University of California Berkeley

  1971