Significance of N-terminal proteolysis of CCL14a to activity on the chemokine receptors CCR1 and CCR5 and the human cytomegalovirus-encoded chemokine receptor US28.
The CC chemokine CCL14a is constitutively expressed in a large variety of tissues and its inactive proform CCL14a(1-74) circulates in high concentrations in plasma. CCL14a(1-74) is converted into CCL14a(9-74) by the proteases urokinase-type plasminogen activator and plasmin and is a highly active agonist for the chemokine receptors CCR1 and CCR5. In this study, a new CCL14a analog, CCL14a(12-74), was isolated from blood filtrate. To elucidate the functional role of the N terminus, a panel of N-terminally truncated CCL14a analogs were tested on the receptors CCR1 to CCR5 and on the human cytomegalovirus (HCMV)-encoded chemokine receptor US28. The rank order of binding affinity to these receptors and of the activation of CCR1 and CCR5-mediated intracellular Ca(2+) concentration mobilization is CCL14a(6-74)<(7-74)<(8-74)<<(9-74) = (10-74)>>(11-74)>>(12-74). The almost identical affinities of CCL14a(7-74), CCL14a(9-74), and CCL14a(10-74) for the US28 receptor and the inhibition of US28-mediated HIV infection of 293T cells by all of the N-terminally truncated CCL14a analogs support the promiscuous nature of the viral chemokine receptor US28. In high concentrations, CCL14a(12-74) did reveal antagonistic activity on intracellular Ca(2+) concentration mobilization in CCR1- and CCR5-transfected cells, which suggests that truncation of Tyr(11) might be of significance for an efficient inactivation of CCL14a. A putative inactivation pathway of CCL14a(9-74) to CCL14a(12-74) may involve the dipeptidase CD26/dipeptidyl peptidase IV (DPPIV), which generates CCL14a(11-74), and the metalloprotease aminopeptidase N (CD13), which displays the capacity to generate CCL14a(12-74) from CCL14a(11-74). Our results suggest that the activity of CCL14a might be regulated by stringent proteolytic activation and inactivation steps.
Richter
Rudolf
R
Institute of Transfusion Medicine and Immune Hematology, Blood Donation Service of the German Red Cross, Frankfurt, Germany. Rudorichter@gmx.de
Casarosa
Paola
P
Ständker
Ludger
L
Münch
Jan
J
Springael
Jean-Yves
JY
Nijmeijer
Saskia
S
Forssmann
Wolf-Georg
WG
Vischer
Henry F
HF
Vakili
Jalal
J
Detheux
Michel
M
Parmentier
Marc
M
Leurs
Rob
R
Smit
Martine J
MJ
D020013:Calcium Signaling; D002460:Cell Line; D019742:Chemokines, CC / Q000378:metabolism*; D003587:Cytomegalovirus; D018819:Dipeptidyl Peptidase 4 / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism; D015658:HIV Infections / Q000517:prevention & control; D006801:Humans; D010446:Peptide Fragments / Q000737:chemistry / Q000502:physiology*; D010447:Peptide Hydrolases / Q000378:metabolism*; D011485:Protein Binding; D054389:Receptors, CCR1 / Q000378:metabolism; D019713:Receptors, CCR5 / Q000378:metabolism; D019707:Receptors, Chemokine / Q000378:metabolism*; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism; D014764:Viral Proteins / Q000378:metabolism
C097720:CCL14 protein, human; D019742:Chemokines, CC; D010446:Peptide Fragments; D054389:Receptors, CCR1; D019713:Receptors, CCR5; D019707:Receptors, Chemokine; C411191:US28 receptor, Cytomegalovirus; D014764:Viral Proteins; D010447:Peptide Hydrolases; D018819:Dipeptidyl Peptidase 4; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator