Abstract
We have cloned and characterized a novel isoform of the skeletal muscle LIM protein 1 (SLIM1), designated SLIMMER. SLIM1 contains an N-terminal single zinc finger followed by four LIM domains. SLIMMER is identical to SLIM1 over the first three LIM domains but contains a novel C-terminal 96 amino acids with three potential bipartite nuclear localization signals, a putative nuclear export sequence, and 27 amino acids identical to the RBP-J binding region of KyoT2, a murine isoform of SLIM1. SLIM1 localized to the cytosol of Sol8 myoblasts and myotubes. SLIMMER was detected in the nucleus of myoblasts and, following differentiation into myotubes, was exclusively cytosolic. Recombinant green fluorescent protein-SLIM1 localized to the cytoplasm and associated with focal adhesions and actin filaments in COS-7 cells, while green fluorescent protein-SLIMMER was predominantly nuclear. SLIMMER truncation mutants revealed that the first nuclear localization signal mediates nuclear localization. The addition of the proposed nuclear export sequence decreased the level of exclusively nuclear expression and increased cytosolic SLIMMER expression in COS-7 cells. The leucine-rich nuclear export signal was required for the export of SLIMMER from the nucleus of myoblasts to the cytoplasm of myotubes. Collectively, these results suggest distinct roles for SLIM1 and SLIMMER in focal adhesions and nuclear-cytoplasmic communication.
Highlights
We have cloned and characterized a novel isoform of the skeletal muscle LIM protein 1 (SLIM1), designated SLIMMER
Sequence analysis revealed that the 1.9-kb cDNA is 99% identical to that of SLIM1 [25], which was derived by analysis of 36 overlapping sequences from a skeletal muscle cDNA library and various sequence data bases, except that our cDNA lacks 400 bp of the 3Ј-untranslated region
Translation of the cDNA sequence reveals that SLIM1 is composed almost entirely of four LIM domains preceded by a single N-terminal zinc finger, with a predicted unmodified molecular mass of 32 kDa
Summary
We have cloned and characterized a novel isoform of the skeletal muscle LIM protein 1 (SLIM1), designated SLIMMER. SLIMMER is identical to SLIM1 over the first three LIM domains but contains a novel C-terminal 96 amino acids with three potential bipartite nuclear localization signals, a putative nuclear export sequence, and 27 amino acids identical to the RBP-J binding region of KyoT2, a murine isoform of SLIM1. The leucine-rich nuclear export signal was required for the export of SLIMMER from the nucleus of myoblasts to the cytoplasm of myotubes These results suggest distinct roles for SLIM1 and SLIMMER in focal adhesions and nuclear-cytoplasmic communication. The LIM domain is a double zinc finger motif that mediates the protein-protein interactions of transcription factors, signaling, and cytoskeleton-associated proteins [1,2,3,4]. Ing pockets stabilizing the tertiary structure of the protein [6, 7] Despite their structural resemblance to GATA-1 zinc fingers, there is no evidence that LIM domains bind DNA directly. An increasing number of studies implicate LIM domains in protein-protein interactions that regulate development, cellular differentiation, and the cytoskeleton [8, 9]
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