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Characterization of heterotrimeric G protein complexes in rice plasma membrane
Author(s) -
Kato Chiyuki,
Mizutani Tomohiro,
Tamaki Hisanori,
Kumagai Hidehiko,
Kamiya Takehiro,
Hirobe Ayumi,
Fujisawa Yukiko,
Kato Hisaharu,
Iwasaki Yukimoto
Publication year - 2004
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1111/j.1365-313x.2004.02046.x
Subject(s) - heterotrimeric g protein , protein subunit , immunoprecipitation , biochemistry , chemistry , yeast , membrane protein , g beta gamma complex , membrane , biology , g alpha subunit , gene , g protein , signal transduction
Summary Two genes in the rice genome were identified as those encoding the γ subunits, γ1 and γ2, of heterotrimeric G proteins. Using antibodies against the recombinant proteins for the α, β, γ1, and γ2 subunits of the G protein complexes, all of the subunits were proven to be localized in the plasma membrane in rice. Gel filtration of solubilized plasma membrane proteins showed that all of the α subunits were present in large protein complexes (about 400 kDa) containing the other subunits, β, γ1, and γ2, and probably also some other proteins, whereas large amounts of the β and γ (γ1 and γ2) subunits were freed from the large complexes and took a 60‐kDa form. A yeast two‐hybrid assay and co‐immunoprecipitation experiments showed that the β subunit interacted tightly with the γ1 and γ2 subunits, and so the β and γ subunits appeared to form dimers in rice cells. Some dimers were associated with the α subunit, because few β, γ1, and γ2 subunits were present in the 400‐kDa complexes in a rice mutant, d1 , which was lacking in the α subunit. When a constitutively active form of the α subunit was prepared by the exchange of one amino acid residue and introduced into d1 , the mutagenized subunit was localized in the plasma membrane of the transformants and took a free, and not the 400‐kDa, form.