Self-aggregation of triadin in the sarcoplasmic reticulum of rabbit skeletal muscle.
The 95 kDa transmembrane glycoprotein triadin is believed to be an essential component of excitation-contraction coupling in the junctional sarcoplasmic reticulum of skeletal muscle fibers. It is debatable whether triadin mediates intraluminal interactions between calsequestrin and the ryanodine receptor exclusively or whether this junctional protein provides also a cytoplasmic linkage between the Ca2+-release channel and the dihydropyridine receptor. Here, we could show that native triadin exists as disulfide-linked homo-polymers of above 3000 kDa. Under non-reducing conditions, protein bands representing the alpha1-dihydropyridine receptor and calsequestrin did not show an immunodecorative overlap with the extremely high-molecular-mass triadin clusters. Following chemical crosslinking, the ryanodine receptor and triadin exhibited a similarly decreased electrophoretic mobility. However, immunoblotting of diagonal non-reducing/reducing two-dimensional gels clearly demonstrated a lack of overlap between the immunodecorated bands representing triadin, the alpha1-dihydropyridine receptor, the ryanodine receptor and calsequestrin. Thus, in native membranes triadin appears to form large self-aggregates primarily. Although triadin exists in a close neighborhood relationship to the Ca2+-release channel tetramers, it does not seem to be directly linked to the other main triad components implicated in the regulation of the excitation-contraction-relaxation cycle and Ca2+-homeostasis. This agrees with a proposed role of triadin in the maintenance of overall triad architecture.
Froemming
G R
GR
Department of Pharmacology, National University of Ireland, University College Dublin, Belfield, Dublin 4, Ireland.
Murray
B E
BE
Ohlendieck
K
K
D000818:Animals; D002352:Carrier Proteins; D015180:Electrophoresis, Gel, Two-Dimensional; D015151:Immunoblotting; D009124:Muscle Proteins / Q000737:chemistry*; D018482:Muscle, Skeletal / Q000737:chemistry / Q000378:metabolism*; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000737:chemistry* / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism
D002352:Carrier Proteins; D009124:Muscle Proteins; C070048:triadin