PLP2/A4 interacts with CCR1 and stimulates migration of CCR1-expressing HOS cells.
Multiple CC chemokines bind to CCR1, which plays important roles in immune and inflammatory responses. To search for proteins involved in the CCR1 signaling pathway, we screened a yeast two-hybrid library using the cytoplasmic tail of CCR1 as the bait. One of the positive clones contained an open reading frame of 456bp, of which the nucleotide sequence was identical to that of proteolipid protein 2 (PLP2), also known as protein A4. Mammalian two-hybrid and coimmunoprecipitation analyses demonstrated the association of PLP2/A4 with CCR1. Indirect immunofluorescence analysis revealed that PLP2/A4 was predominantly located in plasma membrane and colocalized with CCR1 in transfected human HEK293 cells. In addition, focal staining of CCR1 appeared on the periphery of the membrane upon short exposure to Leukotactin-1(Lkn-1)/CCL15, a CCR1 agonist, and was costained with PLP2/A4 on the focal regions. PLP2/A4 mRNAs were detected in various cells such as U-937, HL-60, HEK293, and HOS cells. Overexpression of PLP2/A4 stimulated a twofold increase in the agonist-induced migration of HOS/CCR1 cells, implicating a functional role for PLP2/A4 in the chemotactic processes via CCR1.
Lee
Sang Min
SM
Graduate School of Biotechnology and Institute of Life Sciences and Resources, Kyung Hee University, Yongin, Kyunggi-Do 449-701, Republic of Korea.
Shin
Hwayean
H
Jang
Sung-Wuk
SW
Shim
Jung-Jae
JJ
Song
In-Sung
IS
Son
Kyung-No
KN
Hwang
Jungsu
J
Shin
Yong-Hyun
YH
Kim
Hong-Hee
HH
Lee
Chong-Kil
CK
Ko
Jesang
J
Na
Doe Sun
DS
Kwon
Byoung S
BS
Kim
Jiyoung
J
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D002465:Cell Movement; D002633:Chemotaxis; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D018922:HL-60 Cells; D006801:Humans; D047468:Immunoprecipitation; D062792:MARVEL Domain-Containing Proteins; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011510:Proteolipids; D012313:RNA / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D012509:Sarcoma / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection; D020798:Two-Hybrid System Techniques; D020298:U937 Cells
D062792:MARVEL Domain-Containing Proteins; D008565:Membrane Proteins; C094765:PLP2 protein, human; D011510:Proteolipids; D012313:RNA; D015500:Chloramphenicol O-Acetyltransferase; C071570:SNF1-related protein kinases; D017346:Protein Serine-Threonine Kinases