Mapping of a region within the N terminus of Jak1 involved in cytokine receptor interaction.
Janus kinase 1 (Jak1) is a cytoplasmic tyrosine kinase that noncovalently associates with a variety of cytokine receptors. Here we show that the in vitro translated N-terminal domains of Jak1 are sufficient for binding to a biotinylated peptide comprising the membrane-proximal 73 amino acids of gp130, the signal-transducing receptor chain of interleukin-6-type cytokines. By the fold recognition approach amino acid residues 36-112 of Jak1 were predicted to adopt a beta-grasp fold, and a structural model was built using ubiquitin as a template. Substitution of Tyr(107) to alanine, a residue conserved among Jaks and involved in hydrophobic core interactions of the proposed beta-grasp domain, abrogated binding of full-length Jak1 to gp130 in COS-7 transfectants. By further mutagenesis we identified the loop 4 region of the Jak1 beta-grasp domain as essential for gp130 association and gp130-mediated signal transduction. In Jak1-deficient U4C cells reconstituted with the loop 4 Jak1 mutants L80A/Y81A and Delta(Tyr(81)-Ser(84)), the interferon-gamma, interferon-alpha, and interleukin-6 responses were similarly impaired. Thus, loop 4 of the beta-grasp domain plays a role in the association of Jak1 with both class I and II cytokine receptors. Taken together the structural model and the mutagenesis data provide further insight into the interaction of Janus kinases with cytokine receptors.
Haan
C
C
Department of Biochemistry, Rheinisch Westfälische Technische Hochschule Aachen, Pauwelsstr. 30, 52074 Aachen, Germany.
Is'harc
H
H
Hermanns
H M
HM
Schmitz-Van De Leur
H
H
Kerr
I M
IM
Heinrich
P C
PC
Grötzinger
J
J
Behrmann
I
I
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001665:Binding Sites; D019556:COS Cells; D050822:Cytokine Receptor gp130; D007372:Interferons / Q000494:pharmacology; D053613:Janus Kinase 1; D008562:Membrane Glycoproteins / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018121:Receptors, Cytokine / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014443:Tyrosine / Q000235:genetics / Q000378:metabolism
D015703:Antigens, CD; D008562:Membrane Glycoproteins; D018121:Receptors, Cytokine; D050822:Cytokine Receptor gp130; D014443:Tyrosine; D007372:Interferons; D011505:Protein-Tyrosine Kinases; D053613:Janus Kinase 1; D000409:Alanine