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 ␣II-spectrin binds specifically isoform A of low-molecular-weight phosphotyrosine phosphatase (LMW-PTP). The ␣II-spectrin SH3 domain does not interact with LMW-PTP B or C nor does LMW-PTP A interact with the ␣I-spectrin SH3 domain. The interaction of spectrin with LMW-PTP A led us to look for a tyrosinephosphorylated residue in ␣II-spectrin. Western blotting showed that ␣II-spectrin is tyrosine phosphorylated in vivo. Using mutagenesis on recombinant peptides, we identified the residue Y1176 located in the calpain cleavage site of ␣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.First identified at the intracellular surface of the erythrocyte plasma membrane, spectrins (Sp) are now known to be the central components of the membrane skeleton, a ubiquitous and complex spectrin-actin scaffold located under the lipid bilayer of metazoan animal cells (for review, see references 4 and 21). Numerous studies on red cells, particularly those in hereditary hemolytic anemia, have clearly established the organization of the erythrocyte skeleton and its importance in maintaining erythrocyte shape, stability, and deformability. Spectrins are giant extended flexible molecules composed of two subunits (␣I and ␤I in red cells) which intertwine to form ␣␤ heterodimers. Spectrin exists as elongated tetramers resulting from self-association of ␣␤ heterodimers. Sp tetramers constitute the filaments of the lattice, the nodes of which are crossed-linked by short actin filaments. This spectrin-based skeleton is bound to various transmembrane proteins through two connecting proteins, ankyrin and protein 4.1.In nonerythroid mammal cells, ␣ (␣I and ␣II) and ␤ (␤I to ␤V) chains are encoded by two and five genes, respectively, each of these genes producing several isoforms by alternative splicing. Despite this diversity, all Sp chains present the same structural organization mainly made up of a succession of triple-helical repeat units, 22 for ␣ chains and 17 for ␤ chains except ␤V, which has 30 repeats. These units are characteristic of spectrin family members. They are about 106 amino acids long and folded in a coiled-coil structure made up of three helices (A, B, and C). Beside these repeat units, spectrin isoforms can also con...

Spectrins are ubiquitous scaffolding components of the membrane skeleton that organize and stabilize microdomains on both the plasma membrane and the intracellular organelles. By way of their numerous interactions with diverse protein families, they are implicated in various cellular functions. Using small interfering RNA strategy in the WM-266 cell line derived from human melanoma, we found that ␣II-spectrin deficiency is associated with a defect in cell proliferation, which is related to a cell cycle arrest at the G 1 phase (first gap phase), as evaluated by DNA analysis and Rb phosphorylation. These observations coincided with elevated expression of the cyclin-dependent kinase inhibitor, p21 Cip . Concomitantly, spectrin loss impaired cell adhesion and spreading. These cell adhesion defects were associated with modifications of the actin cytoskeleton, such as loss of stress fibers, alterations of focal adhesions, and modified expression of some integrins. Our results provide novel insights into spectrin functions by demonstrating the involvement of ␣II-spectrin in cell cycle regulation and actin organization.First identified at the intracellular surface of the erythrocyte plasma membrane, the spectrin-based skeleton is considered as a nearly ubiquitous and complex spectrin-actin network in metazoan cells (1).Spectrins are giant extended flexible molecules composed of two subunits (␣ and ␤) that intertwine to form ␣␤ heterodimers. Spectrin is normally considered to exist as tetramers resulting from self-association of ␣␤ dimers. Spectrin tetramers constitute the filaments of the lattice, the nodes of which are cross-linked by actin filaments. This spectrin-based skeleton is bound to various transmembrane proteins either directly, or more frequently through two connecting proteins, ankyrin and protein 4.1. In mammals, the spectrin family currently includes seven genes encoding for two ␣-subunits (␣I and ␣II), four "conventional" ␤-subunits (␤I to ␤IV) and one ␤ heavy subunit (␤V), as well as multiple alternatively spliced variants, each of these species presenting its specific cellular expression pattern. For example, whereas ␣I-spectrin is essentially expressed in the mature erythrocyte, ␣II-spectrin is the most common form in nucleated cells.The functions clearly determined up to date for the spectrin network emerge from human mutations associated with diseases as well as from animal models. Numerous studies on red cells, particularly those in hereditary hemolytic anemia, have clearly established its importance for supporting cell shape and for maintaining cell membrane integrity and stability (2, 3). In nucleated cells, the spectrin-based skeleton has been shown to participate in the stabilization or activation of several specialized membrane proteins, as recently reported for the TRCP channels (4). The direct interaction between TRCP4 channel and spectrin is involved in the regulation of the channel surface expression and activation. One consistent feature observed when spectrin or its binding partner ankyr...