Preparation and characterization of monoclonal antibodies specific for lauroylated isoform of bovine transducin alpha-subunit: immunohistochemical analysis of bovine retinas.
The photoreceptor G protein transducin [alpha- and beta gamma-subunits (T alpha/T beta gamma)] plays a central role in the visual transduction process. The amino-terminus of bovine T alpha is modified by one of four distinct fatty acids-laurate (C12:0), myristate (C14:0), C14:1 (5-cis), and C14:2 (5-cis, 8-cis)-but the biological significance and the localization of the four isoforms of T alpha are poorly understood. To investigate the cellular distribution of each isoform, we prepared monoclonal antibodies against a synthetic C12:0-, C14:0-, C14:1-, or C14:2-nonapeptide corresponding to the N-terminal region of T alpha. Among several types of antibodies isolated, only one type, represented by LA4, reacted specifically with the C12:0-peptide as well as purified T alpha but not with the other proteins in bovine retinal homogenate, including recoverin, indicating that the epitope comprises both C12:0 and the N-terminal amino acids of T alpha. Immunohistochemical analyses of bovine retinal sections by LA4 showed the uniform distribution of C12:0-T alpha in almost all the rod outer segments. Hence, it seemed unlikely that each isoform of T alpha was localized in specific cells. This observation, together with evidence for a possible functional diversity among the isoforms, suggests that the four isoforms of T alpha in a single rod cell may contribute simultaneously to a fine tuning of the photon-signal transduction process.
Shouno
O
O
Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, Japan.
Kokame
K
K
Araki
M
M
Takao
T
T
Shimonishi
Y
Y
Murata
M
M
Yoshizawa
T
T
Fukada
Y
Y
D000215:Acylation; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D002417:Cattle; D007150:Immunohistochemistry; D007536:Isomerism; D010455:Peptides / Q000378:metabolism / Q000494:pharmacology; D012160:Retina / Q000378:metabolism*; D013194:Staining and Labeling; D014018:Tissue Distribution; D015293:Transducin / Q000276:immunology* / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D010455:Peptides; D010566:Virulence Factors, Bordetella; D000246:Adenosine Diphosphate Ribose; D015293:Transducin