Science - USA (2022-04-08)

models for the PK-TK domains that form
head-to-head dimers through interactions
between kinase N lobes, which could poten-
tially sterically occlude substrate binding
and catalytic activity ( 18 , 19 ). Our finding
that activating Val→Phe mutations are posi-
tioned at a central PK-PK dimer interface with-
in the active JAK1-IFNlR1 complex suggest a
simple mechanism for oncogenic activation
in which improved shape complementarity
and hydrophobicity drive ligand-independent
dimerization.
Recent discovery of highly selective TYK2
PK inhibitors, which allosterically stabilize
an autoinhibited conformation underscore
that the JAK family is amenable to devel-
opment of allosteric rather than active-site–
directed inhibitors of kinase function ( 50 , 51 ).
One current challenge in the treatment of
JAK2 V617F patients is resistance to kinase
inhibitors as a result of heterodimerization
and activation of JAK1 and TYK2 ( 52 ). Thus,
new therapies could be designed to directly
target the Val→Phe homodimer interface to
increase specificity and reduce possibility
for escape through activation of other JAKs.
More generally, classification of oncogenic
mutations by their mechanism of action,
either through disruption of autoinhibition or
increased dimerization, may provide a differ-
ential diagnostic criterion to inform therapeu-
tic strategies.
The homodimeric JAK structure visualized
here gives insight into the mechanisms un-
derlying the“tuneability”of cytokine receptor
signaling. Previous studies using genetically
engineered chimeric receptors or engineered
ligands have shown that the geometric varia-
tion of the cytokine receptor dimer can in-
fluence the nature of downstream signaling
( 53 – 56 ). The JAK PK dimer interface could
potentially act as an intracellular fulcrum
in the manner of a ball and socket joint to
reposition the relative orientations and prox-
imities of the C-terminal TK domains resulting
in differential phosphorylation of the recep-
tor ICDs and downstream STATs. In addi-
tion, our structure begins to rationalize how
engineered cytokine ligands can elicit par-
tial agonism. Partial agonists of cytokines
have been engineered through mutational
disruption of the low-affinity“site 2”cytokine
receptor binding site that lowers the effi-
ciency of receptor dimerization ( 57 – 61 ). The
low affinity of the JAK PK dimerization inter-
face might allow small changes in extracellu-
lar affinity to be sensitively transmitted to the
downstream signaling apparatus to regulate
the level of STAT activation. Indeed, the in-
creased affinity of the JAK2 V617F mutant ex-
ploits this natural dimerization interface to
drive ligand-independent signaling.
Many questions remain to refine our under-
standing of the cytokine receptor and JAK

activation process. For example, the confor-

mational transition from the presumed closed

state of the monomeric JAK to the activated

open state in the dimer is largely speculative,

but resolution of this question could provide

new mechanism-based opportunities to mod-

ulate cytokine receptor signaling. The resolu-

tion of the JAK1 homodimeric complex now

allows for the design of small-molecule in-

hibitors of VF dimerization by in silico and ex-

perimental screening approaches based on the

newly resolved PK dimer. Additionally, the

structural basis for how cytokine receptor intra-

cellular domains are phosphorylated at spe-

cific STAT docking sites, followed by binding,

activation, and release of activated phospho-

STATs, are the next frontier in the structural

biology of cytokine receptor signaling.

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ACKNOWLEDGMENTS

We thank R. Fernandes and other members of the Garcia

Laboratory for thoughtful discussion and helpful feedback.

Cryo-EM data were collected at the Stanford cryo-EM center

(cEMc). We thank E. Montabana and Y.-T. Li for generous

support.Funding:This work was supported by National

Institutes of Health grant R37AI51321 (to K.C.G.); Howard

Hughes Medical Institute (to K.C.G.); Ludwig Institute for Cancer

Research (to K.C.G.); Helen Hay Whitney Foundation (to R.A.S.);

National Science Foundation Graduate Research Fellowship

DGE-1656518 (to C.R.G.); Human Frontier Science Program

Organization Fellowship LT000011/2016-L (to N.T.).Author

contributions:Conceptualization: C.R.G., N.T., and K.C.G.

Methodology: K.C.G., C.R.G. and N.T. Investigation: C.R.G.,

N.T., R.A.S., and K.M.J. Funding acquisition: K.C.G. Project

administration: K.C.G. Supervision: K.C.G. Writing–original draft:

K.C.G., C.R.G., N.T., and P.J.L. Writing–review and editing:

K.C.G., C.R.G., N.T., K.M.J., R.A.S., and P.J.L.Competing

interests:K.C.G. is the founder of Synthekine. All other

authors declare no competing interests.Data and materials

availability:The cryo-EM maps have been deposited in the

Electron Microscopy Data Bank (EMDB) under accession code

EMD-25715. The model coordinates have been deposited in the

Protein Data Bank (PDB) under accession code 7T6F.

SUPPLEMENTARY MATERIALS

science.org/doi/10.1126/science.abn8933

Materials and Methods

Figs. S1 to S6

Tables S1 and S2

References ( 62 – 73 )

MDAR Reproducibility Checklist

29 December 2021; accepted 24 February 2022

Published online 10 March 2022

10.1126/science.abn8933

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