TY - JOUR
T1 - Structural Features of an Extended C-Terminal Tail Modulate the Function of the Chemokine CCL21
AU - Moussouras, Natasha A.
AU - Hjortø, Gertrud M.
AU - Peterson, Francis C.
AU - Szpakowska, Martyna
AU - Chevigné, Andy
AU - Rosenkilde, Mette M.
AU - Volkman, Brian F.
AU - Dwinell, Michael B.
N1 - Funding Information:
This work is supported in part by grants from the National Institutes of Health including F30 CA210587 to N.A.M., AI058072 and GM097381 to B.F.V., and U01 CA178960 and R01 CA226279 to M.B.D. Additional support was also provided to M.B.D. by the Advancing a Healthier Wisconsin Endowment and continuing philanthropic support from the Bobbie Nick Voss Charitable Foundation. N.A.M. is a member of the Medical Scientist Training Program at MCW, which is partially supported by a training grant from NIGMS T32-GM080202.
Publisher Copyright:
© 2020 American Chemical Society.
PY - 2020/4/7
Y1 - 2020/4/7
N2 - The chemokines CCL21 and CCL19, through binding of their cognate receptor CCR7, orchestrate lymph node homing of dendritic cells and naïve T cells. CCL21 differs from CCL19 via an unstructured 32 residue C-terminal domain. Previously described roles for the CCL21 C-terminus include GAG-binding, spatial localization to lymphatic vessels, and autoinhibitory modulation of CCR7-mediated chemotaxis. While truncation of the C-terminal tail induced chemical shift changes in the folded chemokine domain, the structural basis for its influence on CCL21 function remains largely unexplored. CCL21 concentration-dependent NMR chemical shifts revealed weak, nonphysiological self-association that mimics the truncation of the C-terminal tail. We generated a series of C-terminal truncation variants to dissect the C-terminus influence on CCL21 structure and receptor activation. Using NMR spectroscopy, we found that CCL21 residues 80-90 mediate contacts with the chemokine domain. In cell-based assays for CCR7 and ACKR4 activation, we also found that residues 92-100 reduced CCL21 potency in calcium flux, cAMP inhibition, and β-arrestin recruitment. Taken together, these structure-function studies support a model wherein intramolecular interactions with specific residues of the flexible C-terminus stabilize a less active monomer conformation of the CCL21. We speculate that the autoinhibitory intramolecular contacts between the C-terminal tail and chemokine body are disrupted by GAG binding and/or interactions with the CCR7 receptor to ensure optimal functionality.
AB - The chemokines CCL21 and CCL19, through binding of their cognate receptor CCR7, orchestrate lymph node homing of dendritic cells and naïve T cells. CCL21 differs from CCL19 via an unstructured 32 residue C-terminal domain. Previously described roles for the CCL21 C-terminus include GAG-binding, spatial localization to lymphatic vessels, and autoinhibitory modulation of CCR7-mediated chemotaxis. While truncation of the C-terminal tail induced chemical shift changes in the folded chemokine domain, the structural basis for its influence on CCL21 function remains largely unexplored. CCL21 concentration-dependent NMR chemical shifts revealed weak, nonphysiological self-association that mimics the truncation of the C-terminal tail. We generated a series of C-terminal truncation variants to dissect the C-terminus influence on CCL21 structure and receptor activation. Using NMR spectroscopy, we found that CCL21 residues 80-90 mediate contacts with the chemokine domain. In cell-based assays for CCR7 and ACKR4 activation, we also found that residues 92-100 reduced CCL21 potency in calcium flux, cAMP inhibition, and β-arrestin recruitment. Taken together, these structure-function studies support a model wherein intramolecular interactions with specific residues of the flexible C-terminus stabilize a less active monomer conformation of the CCL21. We speculate that the autoinhibitory intramolecular contacts between the C-terminal tail and chemokine body are disrupted by GAG binding and/or interactions with the CCR7 receptor to ensure optimal functionality.
UR - http://www.scopus.com/inward/record.url?scp=85083041593&partnerID=8YFLogxK
U2 - 10.1021/acs.biochem.0c00047
DO - 10.1021/acs.biochem.0c00047
M3 - Article
C2 - 32182428
AN - SCOPUS:85083041593
SN - 0006-2960
VL - 59
SP - 1338
EP - 1350
JO - Biochemistry
JF - Biochemistry
IS - 13
ER -