The 30-kD domain of protein 4.1 mediates its binding to the carboxyl terminus of pICln, a protein involved in cellular volume regulation.
Erythrocyte protein 4.1 (P4.1) is an 80-kD cytoskeletal protein that is important for the maintenance of the structural integrity and flexibility of the red blood cell membrane. Limited chymotryptic digestion of erythroid P4.1 yields 4 structural domains corresponding to the 30-, 16-, 10-, and 22/24-kD domains. Using a yeast two-hybrid system, we isolated cDNA clones encoding pICln that specifically interacts with the 30-kD domain of P4.1. In this report, we show that the carboxyl-terminus (amino acid residues 103-237) of pICln binds to the 30-kD domain of P4.1 in a yeast two-hybrid system. The direct association between the 30-kD domain of P4.1 and pICln was further confirmed by the following findings: (1) the S35-methione-labeled pICln specifically bound to both GST/P4.1-80 (80 kD) and GST/P4.1-30 (30 kD) fusion proteins, but not to the proteins that lack the 30-kD domain; (2) coimmunoprecipitation analysis of the cell extracts from transfected SiHa cells showed that pICln and P4.1 associate in transfected cells. It was reported that pICln can form a complex with actin and may play a role involved in cellular volume regulation. The direct association between P4.1 and pICln suggests that pICln may link P4.1-bound cytoskeletal elements to an unidentified volume-sensitive chloride channel.
Tang
C J
CJ
Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan, Republic of China.
Tang
T K
TK
D048429:Cell Size; D018118:Chloride Channels / Q000378:metabolism*; D002712:Chlorides / Q000097:blood; D003598:Cytoskeletal Proteins; D003599:Cytoskeleton / Q000648:ultrastructure; D004912:Erythrocytes / Q000166:cytology*; D006801:Humans; D007473:Ion Channels; D017136:Ion Transport; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D005810:Multigene Family; D009479:Neuropeptides; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017384:Sequence Deletion; D013329:Structure-Activity Relationship; D014162:Transfection
C410035:CLNS1A protein, human; D018118:Chloride Channels; D002712:Chlorides; D003598:Cytoskeletal Proteins; D007473:Ion Channels; D008565:Membrane Proteins; D009479:Neuropeptides; D011993:Recombinant Fusion Proteins; C028841:erythrocyte membrane band 4.1 protein; C120557:erythrocyte membrane protein band 4.1-like 1