Phosphorylation of a new brain-specific septin, G-septin, by cGMP-dependent protein kinase.
The septins are a family of GTPase enzymes, some of which are required for the cytokinesis stage of cell division and others of which are associated with exocytosis. We purified and cloned the cDNA for a 40-kDa protein from rat brain that is a substrate for type I cGMP-dependent protein kinase (PKG). The amino acid sequences of two tryptic peptides of P40 showed high homology to the septins. Molecular cloning revealed the 358-amino acid P40 to be a new member of the septin family. P40 was named G-septin, as it is phosphorylated in vitro by PKG, but relatively poorly by the related cAMP-dependent protein kinase and not by protein kinase C. Two splice variants of G-septin (alpha and beta) were found with distinct N and C termini, but a common GTPase domain. G-septin lacks the C-terminal coiled-coil domain characteristic of all other mammalian septins and uniquely has two predicted phosphorylation site motifs for type I PKG. Photoaffinity labeling with [alpha-(32)P]GTP confirmed that G-septin is a GTP-binding protein. Northern blotting showed that G-septin mRNA (5.0 kilobases) is highly expressed in brain and undetectable in 12 other tissues, indicating that the G-septins are primarily neuronal proteins. Very low levels of 6.0-, 3.4-, and 2.6-kilobase transcripts were found in testis. Our results reveal a new class of brain-specific septins that may be regulated by PKG in neurons.
Xue
J
J
Cell Signalling Unit, Children's Medical Research Institute, Wentworthville 2145, New South Wales, Australia.
Wang
X
X
Malladi
C S
CS
Kinoshita
M
M
Milburn
P J
PJ
Lengyel
I
I
Rostas
J A
JA
Robinson
P J
PJ
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000201:enzymology*; D003001:Cloning, Molecular; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D014644:Genetic Variation; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry; D010766:Phosphorylation; D010802:Phylogeny; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D058112:Septins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D020033:Protein Isoforms; D011994:Recombinant Proteins; D006160:Guanosine Triphosphate; D011494:Protein Kinases; D017869:Cyclic GMP-Dependent Protein Kinases; D020558:GTP Phosphohydrolases; D058112:Septins; C407343:septin 3