Transcripts That Associate with the RNA Binding Protein, DEAD-END (DND1), in Embryonic Stem (ES) Cells

Overview

Journal BMC Mol Biol

Publisher Biomed Central

Specialty Molecular Biology

Date 2011 Aug 20

PMID 21851623

Citations 18

Authors

Rui Zhu

Michelina Iacovino

Elisabeth Mahen

Michael Kyba

Angabin Matin

Affiliations

Soon will be listed here.

Abstract

Background: The RNA binding protein, DEAD END (DND1), is essential for maintaining viable germ cells in vertebrates. It is also a testicular germ cell tumor susceptibility factor in mice. DND1 has been shown to interact with the 3'-untranslated region (3'-UTR) of mRNAs such as P27 and LATS2. Binding of DND1 to the 3'-UTRs of these transcripts blocks the inhibitory function of microRNAs (miRNA) from these transcripts and in this way DND1 helps maintain P27 and LATS2 protein expression. We found that DND1 is also expressed in embryonic stem (ES) cells. Because ES cells share similar gene expression patterns as germ cells, we utilized ES cells to identify additional candidate mRNAs that associate with DND1.

Results: ES cells are readily amenable to genetic modification and easier to culture in vitro compared to germ cells. Therefore, for the purpose of our study, we made a genetically modified, stable, human embryonic stem (hES) cell line that expresses hemagluttinin (HA)-tagged DND1 in a doxycycline (dox) regulatable manner. This line expresses modest levels of HA-DND1 and serves as a good system to study DND1 function in vitro. We used this stable cell line to identify the transcripts that physically interact with DND1. By performing ribonucleoprotein immunoprecipitation (RIP) followed by RT-PCR, we identified that transcripts encoding pluripotency factors (OCT4, SOX2, NANOG, LIN28), cell cycle regulators (TP53, LATS2) and apoptotic factors (BCLX, BAX) are specifically associated with the HA-DND1 ribonucleoprotein complex. Surprisingly, in many cases, bioinformatics analysis of the pulled-down transcripts did not reveal the presence of known DND1 interacting motifs.

Conclusions: Our results indicate that the inducible ES cell line system serves as a suitable in vitro system to identify the mRNA targets of DND1. The RIP-RT results hint at the broad spectrum of mRNA targets that interact with DND1 in ES cells. Based on what is known about DND1 function, our results suggest that DND1 may impose another level of translational regulation to modulate expression of critical factors in ES cells.

Citing Articles

Post-transcriptional repression of CFP-1 expands the regulatory repertoire of LIN-41/TRIM71.

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The RNA binding protein DND1 is elevated in a subpopulation of pro-spermatogonia and targets chromatin modifiers and translational machinery during late gestation.

Ruthig V, Hatkevich T, Hardy J, Friedersdorf M, Mayere C, Nef S PLoS Genet. 2023; 19(3):e1010656.

PMID: 36857387 PMC: 10010562. DOI: 10.1371/journal.pgen.1010656.

Heading towards a dead end: The role of DND1 in germ line differentiation of human iPSCs.

Mall E, Lecanda A, Drexler H, Raz E, Scholer H, Schlatt S PLoS One. 2021; 16(10):e0258427.

PMID: 34653201 PMC: 8519482. DOI: 10.1371/journal.pone.0258427.

The Role of DND1 in Cancers.

Zhang Y, Godavarthi J, Williams-Villalobo A, Polk S, Matin A Cancers (Basel). 2021; 13(15).

PMID: 34359581 PMC: 8345090. DOI: 10.3390/cancers13153679.

A transgenic DND1GFP fusion allele reports in vivo expression and RNA-binding targets in undifferentiated mouse germ cells†.

Ruthig V, Yokonishi T, Friedersdorf M, Batchvarova S, Hardy J, Garness J Biol Reprod. 2021; 104(4):861-874.

PMID: 33394034 PMC: 8324984. DOI: 10.1093/biolre/ioaa233.

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