Z-DNA and its role in RNA polymerase III transcriptional regulation

Active Project Grant Awarded Last action

Award ID
R35GM159821
Recipient
Awarding agency
Obligated
$809,138

Award facts

Assistance listing
93.859 Biomedical Research and Research Training
Completion date 24% complete
Place of performance
Dallas, TX 75390, USA

Award details come from USAspending.gov.

Description

Abstract
DNA can adopt different conformations besides the typical b-form. One such alternative DNA structure is z-DNA which is characterized by unusual biophysical properties. Z-DNA is left handed, does not wrap the histone octamer efficiently, and has two minor grooves. The z-alpha domain, first discovered in ADAR1, specially binds to the z-DNA backbone in a sequence-independent manner. Although z-DNA in the cytoplasm and z-RNA have both described roles in innate immunity, IT is unclear what the function of nuclear z-DNA might be. To map z-DNA in vivo, we used the z-alpha chip-SEQ assay that captures regions of z-DNA in vivo. We found that z- DNA is highly enriched at sites of RNA polymerase III. We suspected that a subunit of RNA POL-III might recognize and induce z-DNA formation. Indeed, POLR3F has cryptic structural homology to the z-alpha domain and POLR3F binds to 79% of all z-DNA sites. Depletion of POLR3F shows that POLR3F is required for z-DNA formation at trna genes. POLR3F depletion also induces expression of nearby RNA polymerase II genes indicating that trna transcription can interfere with neighboring gene expression. In this proposal, we will test whether POLR3F has direct z-DNA binding capacity using biochemical in vitro assays. Next, we will test through separation of function mutants whether z-DNA affinity of POLR3F is required for z-DNA formation and trna transcription by RNA POL-III. We will also evaluate the effect of RNA POL-III transcription on other genes genome wide, as well as its effect on herpes virus transcription. Since missense mutations in POLR3F are found in patients with innate immune defects, we will determine the impact of patient mutations on POLR3F z-DNA binding ability and function. Our integrated proposal will define physiological roles for z-DNA in transcriptional regulation using targeted studies of a previously unrecognized z-DNA binding protein, POLR3F.

Transactions

Action Obligated Date
Amendment $404,569
Original award $404,569

Quick answers

Who received award R35GM159821?

R35GM159821 was awarded to The University Of Texas Southwestern Medical Center by National Institute of General Medical Sciences (National Institutes of Health) on September 1, 2025.

How much is award R35GM159821 worth?

$809,138 has been obligated on R35GM159821.

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