Tyrosine phosphorylation regulates alpha II spectrin cleavage by calpain.
Spectrins, components of the membrane skeleton, are implicated in various cellular functions. Understanding the diversity of these functions requires better characterization of the interacting domains of spectrins, such as the SH3 domain. Yeast two-hybrid screening of a kidney cDNA library revealed that the SH3 domain of alpha II-spectrin binds specifically isoform A of low-molecular-weight phosphotyrosine phosphatase (LMW-PTP). The alpha II-spectrin SH3 domain does not interact with LMW-PTP B or C nor does LMW-PTP A interact with the alpha I-spectrin SH3 domain. The interaction of spectrin with LMW-PTP A led us to look for a tyrosine-phosphorylated residue in alpha II-spectrin. Western blotting showed that alpha II-spectrin is tyrosine phosphorylated in vivo. Using mutagenesis on recombinant peptides, we identified the residue Y1176 located in the calpain cleavage site of alpha II-spectrin, near the SH3 domain, as an in vitro substrate for Src kinase and LMW-PTP A. This Y1176 residue is also an in vivo target for kinases and phosphatases in COS cells. Phosphorylation of this residue decreases spectrin sensitivity to calpain in vitro. Similarly, the presence of phosphatase inhibitors in cell culture is associated with the absence of spectrin cleavage products. This suggests that the Y1176 phosphorylation state could modulate spectrin cleavage by calpain and may play an important role during membrane skeleton remodeling.
Nicolas
Gaël
G
INSERM U409, Faculté de Médecine Xavier Bichat-Association Claude Bernard, Paris, France.
Fournier
Catherine M
CM
Galand
Colette
C
Malbert-Colas
Laurence
L
Bournier
Odile
O
Kroviarski
Yolande
Y
Bourgeois
Monique
M
Camonis
Jacques H
JH
Dhermy
Didier
D
Grandchamp
Bernard
B
Lecomte
Marie-Christine
MC
D000595:Amino Acid Sequence; D000818:Animals; D002154:Calpain / Q000378:metabolism*; D002458:Cell Fractionation; D002460:Cell Line; D003599:Cytoskeleton / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D010766:Phosphorylation; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D016415:Sequence Alignment; D013049:Spectrin / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014443:Tyrosine / Q000378:metabolism*; D015003:Yeasts / Q000235:genetics / Q000378:metabolism
D020033:Protein Isoforms; D013049:Spectrin; D014443:Tyrosine; D017027:Protein Tyrosine Phosphatases; D002154:Calpain; C449002:mu-calpain