Structural analysis and tissue localization of human C4.4A: a protein homologue of the urokinase receptor.
C4.4A, a structural homologue of the urokinase-type plasminogen activator receptor (uPAR), was originally identified as a metastasis-associated membrane protein, but little is known about its structural and functional properties. Therefore, we expressed, purified and characterized a soluble truncated form of human C4.4A, and used this protein to produce specific polyclonal anti-C4.4A antibodies. By immunohistochemistry we observed a pronounced surface staining for C4.4A in suprabasal keratinocytes of chronic human wounds and found C4.4A expression markedly upregulated in migrating keratinocytes during re-epithelisation of incisional skin wounds. Phorbol-ester-induced hyperplasia of mouse skin is also accompanied by a significant induction of C4.4A expression in the multilayered, suprabasal keratinocytes. C4.4A contains two Ly-6 (leucocyte antigen 6)/uPAR/alpha-neurotoxin modules. Our recombinant human C4.4A is extensively modified by post-translational glycosylation, which include 5-6 N-linked carbohydrates primarily located in or close to its second Ly-6/uPAR/alpha-neurotoxin module and approximately 15 O-linked carbohydrates clustered in a Ser/Thr/Pro-rich region at the C-terminus. A highly protease-sensitive region (Tyr200-Arg204) is located between these two clusters of N- and O-linked carbohydrates. The natural, glycolipid-anchored C4.4A from amnion membranes of human term placenta exhibits similar properties. Using recombinant, soluble C4.4A or MCF 7 cells, which express significant amounts of GPI-anchored C4.4A, we find no evidence for an interaction between C4.4A and uPA, a property suggested previously for rat C4.4A. Collectively these data indicate that C4.4A, although being a structural homologue of uPAR, is unlikely to have a functional overlap with uPAR.
Hansen
Line V
LV
Finsen Laboratory, Rigshospitalet, Strandboulevarden 49, DK-2100 Copenhagen, Denmark.
Gårdsvoll
Henrik
H
Nielsen
Boye S
BS
Lund
Leif R
LR
Danø
Keld
K
Jensen
Ole N
ON
Ploug
Michael
M
D000595:Amino Acid Sequence; D000818:Animals; D001943:Breast Neoplasms / Q000473:pathology; D015815:Cell Adhesion Molecules / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D062109:Dermatologic Surgical Procedures; D058851:GPI-Linked Proteins; D006801:Humans; D006965:Hyperplasia / Q000139:chemically induced / Q000378:metabolism; D007150:Immunohistochemistry; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010449:Peptide Mapping / Q000379:methods; D010920:Placenta / Q000737:chemistry; D025941:Protein Interaction Mapping / Q000379:methods; D011499:Protein Processing, Post-Translational / Q000235:genetics; D011956:Receptors, Cell Surface / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D055293:Receptors, Urokinase Plasminogen Activator; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D012867:Skin / Q000737:chemistry / Q000473:pathology; D013755:Tetradecanoylphorbol Acetate / Q000009:adverse effects / Q000031:analogs & derivatives*; D014945:Wound Healing
D015815:Cell Adhesion Molecules; D058851:GPI-Linked Proteins; C472589:LYPD3 protein, human; C524898:PLAUR protein, human; D010446:Peptide Fragments; C524899:Plaur protein, mouse; C524900:Plaur protein, rat; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D011993:Recombinant Fusion Proteins; C016658:phorbolol myristate acetate; D013755:Tetradecanoylphorbol Acetate