Abstract

Paramecium offers advantages in the study of several animal phenomena and is well suited for the genetic dissection of bioelectricity and cell behavior. Over the last 30 years, mutants with defective Ca2+ currents, K+ currents and Na+ currents have been isolated and characterized phenotypically. We have recently developed a method to systematically clone Paramecium genes that correspond to phenotypes. This method entails phenotypic complementation and guarantees the genes cloned to be functional in their biological context. We plan to use this method to harvest the genes from the above mutants. We have cloned the pwA and pwB genes that control the ciliary Ca2+ currents. Based on features of the predicated gene products, we will continue to test their biochemical functions. These and other gene products may give insights to signal transductions and Ca2+ metabolism in cilia and sperm tail. Genes have also been cloned by their homology to known genes of other organisms. We will continue to find ways to create live Paramecium mutants with these homologous genes knocked out. Three methods will be tried: gene silencing, kimeraplasty, and the manipulation of internal eliminated sequences. Because a convenient transformation method with facilitate research of the whole community, we also plan to perfect methods of transforming paramecia en masse.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM022714-28
Application #
6518927
Study Section
Genetics Study Section (GEN)
Program Officer
Shapiro, Bert I
Project Start
1995-05-15
Project End
2003-04-30
Budget Start
2002-05-01
Budget End
2003-04-30
Support Year
28
Fiscal Year
2002
Total Cost
$248,838
Indirect Cost

  Institution

Name
University of Wisconsin Madison
Department
Biochemistry
Type
Other Domestic Higher Education
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715

  Publications

Haynes, W John; Ling, Kit-Yin; Saimi, Yoshiro et al. (2003) PAK paradox: Paramecium appears to have more K(+)-channel genes than humans. Eukaryot Cell 2:737-45
Haynes, W John; Kung, Ching; Saimi, Yoshiro et al. (2002) An exchanger-like protein underlies the large Mg2+ current in Paramecium. Proc Natl Acad Sci U S A 99:15717-22
Ling, K Y; Haynes, W J; Oesterle, L et al. (2001) K(+)-channel transgenes reduce K(+) currents in Paramecium, probably by a post-translational mechanism. Genetics 159:987-95
Hauser, K; Haynes, W J; Kung, C et al. (2000) Expression of the green fluorescent protein in Paramecium tetraurelia. Eur J Cell Biol 79:144-9
Haynes, W J; Ling, K Y; Preston, R R et al. (2000) The cloning and molecular analysis of pawn-B in Paramecium tetraurelia. Genetics 155:1105-17
Kung, C; Saimi, Y; Haynes, W J et al. (2000) Recent advances in the molecular genetics of Paramecium. J Eukaryot Microbiol 47:4-Nov
Loukin, S H; Saimi, Y (1999) K(+)-dependent composite gating of the yeast K(+) channel, Tok1. Biophys J 77:3060-70
Chan, C W; Saimi, Y; Kung, C (1999) A new multigene family encoding calcium-dependent calmodulin-binding membrane proteins of Paramecium tetraurelia. Gene 231:21-32
Haynes, W J; Vaillant, B; Preston, R R et al. (1998) The cloning by complementation of the pawn-A gene in Paramecium. Genetics 149:947-57
Loukin, S H; Vaillant, B; Zhou, X L et al. (1997) Random mutagenesis reveals a region important for gating of the yeast K+ channel Ykc1. EMBO J 16:4817-25

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