Identification and characterization of a novel sucrose-non-fermenting protein kinase/AMP-activated protein kinase-related protein kinase, SNARK.
Subtraction hybridization after the exposure of keratinocytes to ultraviolet radiation identified a differentially expressed cDNA that encodes a protein of 630 amino acid residues possessing significant similarity to the catalytic domain of the sucrose-non-fermenting protein kinase (SNF1)/AMP-activated protein kinase (AMPK) family of serine/threonine protein kinases. Northern blotting and reverse-transcriptase-mediated PCR demonstrated that mRNA transcripts for the SNF1/AMPK-related kinase (SNARK) were widely expressed in rodent tissues. The SNARK gene was localized to human chromosome 1q32 by fluorescent in situ hybridization. SNARK was translated in vitro to yield a single protein band of approx. 76 kDa; Western analysis of transfected baby hamster kidney (BHK) cells detected two SNARK-immunoreactive bands of approx. 76-80 kDa. SNARK was capable of autophosphorylation in vitro; immunoprecipitated SNARK exhibited phosphotransferase activity with the synthetic peptide substrate HMRSAMSGLHLVKRR (SAMS) as a kinase substrate. SNARK activity was significantly increased by AMP and 5-amino-4-imidazolecarboxamide riboside (AICAriboside) in rat keratinocyte cells, implying that SNARK might be activated by an AMPK kinase-dependent pathway. Furthermore, glucose deprivation increased SNARK activity 3-fold in BHK fibroblasts. These findings identify SNARK as a glucose- and AICAriboside-regulated member of the AMPK-related gene family that represents a new candidate mediator of the cellular response to metabolic stress.
Lefebvre
D L
DL
Department of Medicine, Division of Dermatology, Toronto General Hospital, Banting Institute, Room 317, 100 College Street, Toronto M5G 1L5, Ontario, Canada.
Bai
Y
Y
Shahmolky
N
N
Sharma
M
M
Poon
R
R
Drucker
D J
DJ
Rosen
C F
CF
D000249:Adenosine Monophosphate / Q000494:pharmacology; D000595:Amino Acid Sequence; D000620:Aminoimidazole Carboxamide / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D002874:Chromosome Mapping; D006224:Cricetinae; D018076:DNA, Complementary; D004789:Enzyme Activation; D005947:Glucose / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011233:Precipitin Tests; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D012265:Ribonucleotides / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D018076:DNA, Complementary; D012333:RNA, Messenger; D012265:Ribonucleotides; D000620:Aminoimidazole Carboxamide; D000249:Adenosine Monophosphate; C426226:Nuak2 protein, rat; C486061:NUAK2 protein, human; D017346:Protein Serine-Threonine Kinases; C031143:AICA ribonucleotide; D005947:Glucose