Jak2 tyrosine kinase residues glutamic acid 1024 and arginine 1113 form a hydrogen bond interaction that is essential for Jak-STAT signal transduction.
Angiotensin II is a well-known vasoactive peptide, but it can also act as a potent growth factor, partially through activation of the tyrosine kinase Jak2. Activated Jak2 tyrosine phosphorylates and activates members of the Signal Transducers and Activators of Transcription (STAT) family of cytoplasmic transcription factors. Recently, we demonstrated that tryptophan 1020 and glutamic acid 1024 within the Jak2 activation loop are required for Jak2 tyrosine kinase activity. Here, we sought to elucidate the requirement of glutamic acid 1024 for Jak2 function. Using molecular modeling algorithms of the Jak2 kinase domain, we identified a putative interaction between glutamic acid 1024 and an arginine at position 1113. We generated a series of charge-based substitution mutations at position 1113 and found that conversion of arginine 1113 to glutamic acid, alanine, or lysine prevented Jak2 autophosphorylation. Furthermore, mutation of arginine 1113 prevented the following angiotensin II-dependent processes from occurring: (1) Jak2 tyrosine phosphorylation, (2) Jak2/AT1receptor co-association, (3) STAT1 recruitment to the Jak2/AT1receptor complex, (4) STAT1 tyrosine phosphorylation, and (5) STAT-mediated gene expression. We determined that the interaction between glutamic acid 1024 and arginine 1113 consists of two distinct hydrogen bonds. We conclude that these hydrogen bond interactions are critical for Jak2 kinase function and subsequent angiotensin II-dependent activation of the Jak/STAT signaling pathway.
Sandberg
Eric M
EM
Department of Physiology and Functional Genomics, University of Florida College of Medicine, PO Box 100274, Gainesville, FL 32610, USA.
VonDerLinden
Dannielle
D
Ostrov
David A
DA
Sayeski
Peter P
PP
D000409:Alanine / Q000737:chemistry; D000465:Algorithms; D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000737:chemistry; D000818:Animals; D001120:Arginine / Q000737:chemistry*; D015153:Blotting, Western; D019556:COS Cells; D002460:Cell Line; D018360:Crystallography, X-Ray; D003593:Cytoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D005799:Genes, Dominant; D018698:Glutamic Acid / Q000737:chemistry*; D006860:Hydrogen Bonding; D047468:Immunoprecipitation; D053614:Janus Kinase 2; D008156:Luciferases / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D050794:STAT1 Transcription Factor; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D015534:Trans-Activators / Q000378:metabolism*; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry
D004268:DNA-Binding Proteins; D011518:Proto-Oncogene Proteins; D050794:STAT1 Transcription Factor; D015534:Trans-Activators; D000804:Angiotensin II; D018698:Glutamic Acid; D014443:Tyrosine; D001120:Arginine; D008156:Luciferases; D011505:Protein-Tyrosine Kinases; D053614:Janus Kinase 2; D008239:Lysine; D000409:Alanine