TheaimofourproposalistoelucidatethemechanismsthatlinktranscriptionofspecificRNAsinthe nucleustotheirtranslation(RNAprocessing).?Wehaveidentifiedandcharacterizedanovelandhighly conservedgene,?Zfrp8/PDCD2,?andshownthatitisessentialinstemcellsinflies,mouse,andhuman,and thatitisalsorequiredforgrowthofcancercells.Wediscovered?Zfrp8/PDCD2?isrequiredforthenuclearexport ofselectmRNAsandTEtranscripts.Alsoitinteractswiththesmallribosomalsubunitandformsacomplex withmRNAbindingproteins.OurdatasuggestthatZfrp8/PDCD2controlssubcellularlocalizationofselect RNAsandtheassociationofmRNARNPswithribosomes.WealsohaveidentifiedTet/TET1asanew Zfrp8/PDCD2interactingprotein.Invertebrates,TETproteinsfunctioninDNAdemethylationconverting 5methylcytosine(5mC)into5hydroxymethylcytosine(5hmC),modificationsthatarenotdetectedin DrosophilaDNA.ArecentstudyshowsthatvertebrateTetproteinscanalsoconvert5mrCto5hmrCinRNA. Inspiredbythisdiscovery,wehaveshownthat5hmrCalsoexistsinfliesanddependsonTetactivity.We hypothesizethatTetmodifiesspecifictranscriptsandregulatestherecruitmentofZfrp8totheseRNAs,so controllingtheirprocessingandtranslation. Weproposetotestthishypothesisbyacombinationofmolecular/biochemicalandgenetic experiments.Wewillidentify5hmrCmodifiedtranscriptstranscriptomewideandstudytheirintegrity,levels, andlocalizationinwildtypeandmutanttissues.WewillmaptheTetbindingsitesonDNAandcomparetheir locationto5hmrCmodifiedtranscripts.Finally,wewilltesthow?Tet?and?Zfrp8?affectribosomaloccupancyof mRNAsandestablishhow?Tet?and5hmrCaffecttheirtranslation.Inadditiontoinvestigatingthemechanismby which?Tet?and5hmrCregulateRNAmetabolismandhowZ?frp8?affectstheprocess,wewilldeterminethe importanceofbothgenesindevelopmentandstemcelldifferentiationbystudyingthemutantphenotypesof new?Tet?allelesand?Tet?and?Zfrp8KD?tissues.BecauseoftheconservationofbothTetandZfrp8/PDCD2our resultsarelikelytoshedlightonthesameprocessinmouseandhuman.

Public Health Relevance

We propose a study of the link between the transcription of the RNA in the nucleus and the mechanism that selects some of these RNAs for protein translation in the cytoplasm. We study two proteins that control these steps. Absence of these proteins results in an arrest of cell divisions in stem cells, and is also associated with infertility, neurological defects, and cancer.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM120405-04
Application #
9698374
Study Section
Development - 1 Study Section (DEV1)
Program Officer
Reddy, Michael K
Project Start
2016-09-01
Project End
2020-05-31
Budget Start
2019-06-01
Budget End
2020-05-31
Support Year
4
Fiscal Year
2019
Total Cost
Indirect Cost
Name
Rutgers University
Department
Type
Organized Research Units
DUNS #
001912864
City
Piscataway
State
NJ
Country
United States
Zip Code
08854
Wang, Fei; Minakhina, Svetlana; Tran, Hiep et al. (2018) Tet protein function during Drosophila development. PLoS One 13:e0190367