The nuclear lamina, a filamentous protein meshwork lining the nuclear envelope (NE), contains a polymer of lamins and associated transmembrane proteins of the inner nuclear membrane. The importance of the lamina in cellular functions is underscored by the findings that mutations in the genes for lamins A/C and lamina-associated transmembrane proteins cause over 15 inherited human disorders (""""""""laminopathies""""""""), including many dystrophies that affect heart and skeletal muscle. The long-term objectives of this project are to promote an understanding of the lamina's role in cell organization and functions, and its involvement in human diseases. Recently, 67 novel putative NE transmembrane proteins (NETs) have been identified in a comprehensive proteomics analysis of the NE. Expression profiling has revealed that 6 of these NETs are significantly up-regulated during myoblast differentiation and are expressed at high levels in adult skeletal muscle relative to other tissues, indicating an important role in muscle development. This proposal will focus on a 3 of these NETs, which have been validated to be authentic NE proteins, and which have putative roles in signaling based on their homologies to known proteins.
The specific aims are to: 1) Carry out RNAi- mediated gene silencing in a myoblast differentiation model, and molecularly analyze the phenotypic defects that may arise related to differentiation, signaling responses related to muscle regulation, or NE structure. 2) Biochemically analyze specific NET functions suggested by their sequence homologies, and immunolocalize the endogenous NETs at the light and EM levels. 3) Implement MudPIT proteomic analysis of NET- containing protein complexes solubilized from cells to identify other NE proteins that interact with the NETs, and use in vitro binding studies to extend the results of the proteomics. This work is expected to provide a new level of insight on the NE as a signaling platform in the nucleus, and to promote an understanding of how mutations in NE and lamina proteins cause human diseases, particularly diseases affecting striated muscle.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM028521-27
Application #
7541333
Study Section
Nuclear Dynamics and Transport (NDT)
Program Officer
Shapiro, Bert I
Project Start
1988-04-01
Project End
2010-12-31
Budget Start
2009-01-01
Budget End
2009-12-31
Support Year
27
Fiscal Year
2009
Total Cost
$385,917
Indirect Cost
Name
Scripps Research Institute
Department
Type
DUNS #
781613492
City
La Jolla
State
CA
Country
United States
Zip Code
92037
Gerace, Larry; Tapia, Olga (2018) Messages from the voices within: regulation of signaling by proteins of the nuclear lamina. Curr Opin Cell Biol 52:14-21
Stroud, Matthew J; Fang, Xi; Zhang, Jianlin et al. (2018) Luma is not essential for murine cardiac development and function. Cardiovasc Res 114:378-388
Stroud, Matthew J; Feng, Wei; Zhang, Jianlin et al. (2017) Nesprin 1?2 is essential for mouse postnatal viability and nuclear positioning in skeletal muscle. J Cell Biol 216:1915-1924
Tapia, Olga; Gerace, Larry (2016) Analysis of Nuclear Lamina Proteins in Myoblast Differentiation by Functional Complementation. Methods Mol Biol 1411:177-94
Tapia, Olga; Fong, Loren G; Huber, Michael D et al. (2015) Nuclear envelope protein Lem2 is required for mouse development and regulates MAP and AKT kinases. PLoS One 10:e0116196
Huber, Michael D; Vesely, Paul W; Datta, Kaustuv et al. (2013) Erlins restrict SREBP activation in the ER and regulate cellular cholesterol homeostasis. J Cell Biol 203:427-36
Kerkow, Donald E; Carmel, Andrew B; Menichelli, Elena et al. (2012) The structure of the NXF2/NXT1 heterodimeric complex reveals the combined specificity and versatility of the NTF2-like fold. J Mol Biol 415:649-65
Gerace, Larry; Huber, Michael D (2012) Nuclear lamina at the crossroads of the cytoplasm and nucleus. J Struct Biol 177:24-31
Hintersteiner, Martin; Ambrus, Géza; Bednenko, Janna et al. (2010) Identification of a small molecule inhibitor of importin ? mediated nuclear import by confocal on-bead screening of tagged one-bead one-compound libraries. ACS Chem Biol 5:967-79
Ambrus, Geza; Whitby, Landon R; Singer, Eric L et al. (2010) Small molecule peptidomimetic inhibitors of importin ýý/ýý mediated nuclear transport. Bioorg Med Chem 18:7611-20

Showing the most recent 10 out of 32 publications