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Identification of Residues of the Caenorhabditis elegans LIN-1 ETS Domain That Are Necessary for DNA Binding and Regulation of Vulval Cell Fates

Miley, Ginger R. and Fantz, Douglas and Glossip, Danielle and Lu, Xiaowei and Saito, R. Mako and Palmer, Robert E. and Inoue, Takao and van den Heuvel, Sander and Sternberg, Paul W. and Kornfeld, Kerry (2004) Identification of Residues of the Caenorhabditis elegans LIN-1 ETS Domain That Are Necessary for DNA Binding and Regulation of Vulval Cell Fates. Genetics, 167 (4). pp. 1697-1709. ISSN 0016-6731. PMCID PMC1471005. doi:10.1534/genetics.104.029017.

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LIN-1 is an ETS domain protein. A receptor tyrosine kinase/Ras/mitogen-activated protein kinase signaling pathway regulates LIN-1 in the P6.p cell to induce the primary vulval cell fate during Caenorhabditis elegans development. We identified 23 lin-1 loss-of-function mutations by conducting several genetic screens. We characterized the molecular lesions in these lin-1 alleles and in several previously identified lin-1 alleles. Nine missense mutations and 10 nonsense mutations were identified. All of these lin-1 missense mutations affect highly conserved residues in the ETS domain. These missense mutations can be arranged in an allelic series; the strongest mutations eliminate most or all lin-1 functions, and the weakest mutation partially reduces lin-1 function. An electrophoretic mobility shift assay was used to demonstrate that purified LIN-1 protein has sequence-specific DNA-binding activity that required the core sequence GGAA. LIN-1 mutant proteins containing the missense substitutions had dramatically reduced DNA binding. These experiments identify eight highly conserved residues of the ETS domain that are necessary for DNA binding. The identification of multiple mutations that reduce the function of lin-1 as an inhibitor of the primary vulval cell fate and also reduce DNA binding suggest that DNA binding is essential for LIN-1 function in an animal.

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Sternberg, Paul W.0000-0002-7699-0173
Additional Information:© 2004 by the Genetics Society of America. Manuscript received April 7, 2004; Accepted for publication May 6, 2004. We are grateful to the following individuals for providing lin-1 alleles: Bob Horvitz and members of his lab including Greg Beitel, Jeff Thomas, Chip Ferguson, Erik Jorgenson, Scott Clark, Craig Ceol, Frank Stegmeier, Melissa Harrison, and Na An as well as Joseph Lee, Dave Eisenmann, and Stuart Kim. We thank Parie Garg and Andrew Turk for assistance with experiments. Some strains were provided by the Caenorhabditis Genetics Center, which is funded by the National Center for Research Resources of the National Institutes of Health (NIH). This work was supported by grants from the NIH to K.K. (CA-84271). K.K. is a scholar of the Leukemia and Lymphoma Society. P.W.S is an Investigator and R.E.P. was an associate with the Howard Hughes Medical Institute. D.F. was supported by an NIH grant (GM20450-01). R.M.S. has a postdoctoral fellowship from the American Cancer Society and the Massachusetts General Hospital Fund for Medical Discovery.
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Leukemia and Lymphoma SocietyUNSPECIFIED
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
American Cancer SocietyUNSPECIFIED
Massachusetts General HospitalUNSPECIFIED
Issue or Number:4
PubMed Central ID:PMC1471005
Record Number:CaltechAUTHORS:20190514-101613052
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Official Citation:Identification of Residues of the Caenorhabditis elegans LIN-1 ETS Domain That Are Necessary for DNA Binding and Regulation of Vulval Cell Fates. Ginger R. Miley, Douglas Fantz, Danielle Glossip, Xiaowei Lu, R. Mako Saito, Robert E. Palmer, Takao Inoue, Sander van den Heuvel, Paul W. Sternberg and Kerry Kornfeld. GENETICS August 1, 2004 vol. 167 no. 4 1697-1709;
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:95478
Deposited By: Tony Diaz
Deposited On:14 May 2019 20:55
Last Modified:16 Nov 2021 17:13

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