Caenorhabditis elegans TBX-2 Directly Regulates.pdf (1.51 MB)
Caenorhabditis elegans TBX-2 Directly Regulates Its Own Expression in a Negative Autoregulatory Loop
journal contributionposted on 2016-08-29, 00:00 authored by Angenee C. Milton, Peter G. Okkema
T-box genes often exhibit dynamic expression patterns, and their expression levels can be crucial for normal function. Despite the importance of these genes, there is little known about T-box gene regulation. We have focused on the Caenorhabditis elegans gene tbx-2 to understand how T-box gene expression is regulated, and here we demonstrate TBX-2 itself directly represses its own expression in a negative auto-regulatory loop. tbx-2 is essential for normal pharyngeal muscle development, and a tbx-2 promoter gfp fusion (Ptbx-2::gfp) is transiently expressed in the pharynx during embryogenesis and in a small number of head neurons in larvae and adults. Reduced tbx-2 function resulted in ectopic Ptbx-2::gfp expression in the seam cells and gut in larvae and adults. Mutation of potential T-box binding sites within the tbx-2 promoter resulted in a similar pattern of ectopic Ptbx-2::gfp expression, and chromatin immunoprecipitation analyses show TBX-2 binds these sites in vivo. This pattern of ectopic Ptbx-2::gfp expression in tbx-2 mutants was very similar to that observed in mutants affecting the NF-Y complex, and our results comparing tbx-2 and nfyb-1 single- and double mutants suggest TBX-2 and NF-Y function in a single pathway to repress the tbx-2 promoter. The tbx-2 promoter is the first direct target identified for TBX-2, and we used it to ask whether SUMOylation is essential for TBX-2 repression. RNAi knockdown of SUMOylation pathway components led to ectopic Ptbx-2::gfp expression in the seam cells and gut. Ectopic Ptbx-2::gfp also was observed in the syncytial hypodermis, suggesting either the tbx-2 promoter is repressed by other SUMOylation dependent mechanisms, or that decreased SUMOylation leads to stable changes in seam cell nuclei as they fuse with the syncytial hypodermis. We suggest negative autoregulation is an important mechanism that allows precise control of tbx-2 expression levels and may allow rapid changes in gene expression during development.
This project was supported by National Institutes of Health (NIH) grant R01 GM82865 (P.G.O.), an NIH Kirschstein NRSA F31 GM090675 (A.C.M.), and a UIC LAS Award for Faculty in the Natural Sciences. Some strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440).Research Open Access Publishing (ROAAP) Fund of the University of Illinois at Chicago for financial support towards the open access publishing fee for this article.
Publisher StatementThis is a copy of an article published in the G3. © 2015 Milton and Okkema. http://www.g3journal.org/content/early/2015/04/14/g3.115.018101
PublisherGenetics Society of America