The hydrophilic domain of small ankyrin-1 interacts with the two N-terminal immunoglobulin domains of titin.
Little is known about the mechanisms that organize the internal membrane systems in eukaryotic cells. We are addressing this question in striated muscle, which contains two novel systems of internal membranes, the transverse tubules and the sarcoplasmic reticulum (SR). Small ankyrin-1 (sAnk1) is an approximately 17-kDa transmembrane protein of the SR that concentrates around the Z-disks and M-lines of each sarcomere. We used the yeast two-hybrid assay to determine whether sAnk1 interacts with titin, a giant myofibrillar protein that organizes the sarcomere. We found that the hydrophilic cytoplasmic domain of sAnk1 interacted with the two most N-terminal Ig domains of titin, ZIg1 and ZIg2, which are present at the Z-line in situ. Both ZIg1 and ZIg2 were required for binding activity. sAnk1 did not interact with other sequences of titin that span the Z-disk or with Ig domains of titin near the M-line. Titin ZIg1/2 also bound T-cap/telethonin, a 19-kDa protein of the Z-line. We show that titin ZIg1/2 could form a three-way complex with sAnk1 and T-cap. Our results indicate that titin ZIg1/2 can bind sAnk1 in muscle homogenates and suggest a role for these proteins in organizing the SR around the contractile apparatus at the Z-line.
Kontrogianni-Konstantopoulos
Aikaterini
A
Department of Physiology, University of Maryland School of Medicine, Baltimore 21201, USA. akons001@umaryland.edu
Bloch
Robert J
RJ
D017487:Ankyrins / Q000378:metabolism*; D001483:Base Sequence; D001665:Binding Sites; D064211:Connectin; D017931:DNA Primers; D006801:Humans; D007136:Immunoglobulins / Q000378:metabolism*; D009124:Muscle Proteins / Q000737:chemistry / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism; D011485:Protein Binding; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
C110202:ANK1 protein, human; D017487:Ankyrins; D064211:Connectin; D017931:DNA Primers; D007136:Immunoglobulins; D009124:Muscle Proteins; C577285:TTN protein, human; D011494:Protein Kinases