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Muscle-specific mRNA isoform encodes a protein composed mainly of the N-terminal 175 residues of type 2 Ins(1,4,5)P3
A Futatsugi1, G Kuwajima, K Mikoshiba
1CNS Research Laboratories, Shionogi & Co. Ltd., 2-5-1 Mishima, Settsu-shi, Osaka 566-0022, Japan. Shirokanedai, Minato-ku, Tokyo 108-8639,
Abstract:
We have found a novel isoform of the mouse type 2 Ins(1,4,5)P3 receptor [Ins(1,4,5)P3R] mRNA by reverse transcriptase-mediated PCR analysis. The novel isoform, which was expressed specifically in skeletal muscle and heart, was generated by the inclusion of a novel exon. As this exon contains a stop codon, the isoform encodes a putative protein (designated TIPR) consisting of 175 acid residues of the type 2 Ins(1,4,5)P3R and the following six residues derived from this exon. We transfected the cDNA of this isoform into COS-7 cells; these cells expressed a 24 kDa protein that was recognized by an antibody against TIPR produced in Escherichia coli. The isoform encoding TIPR was also found in human skeletal muscle and heart. The N-terminal region of Ins(1,4,5)P3R is suggested to have a role in ligand binding and to interact with the C-terminal channel domain of Ins(1,4,5)P3R itself. TIPR might regulate the Ins(1,4,5)P3 signal pathway in both muscles.
Insights
Researchers discovered a new type 2 Ins(1,4,5)P3 receptor (Ins(1,4,5)P3R) mRNA isoform in mouse and human muscles. This novel isoform produces a unique protein, TIPR, potentially regulating the Ins(1,4,5)P3 signaling pathway.
Area of Science:
- Molecular Biology
- Biochemistry
- Cell Biology
Background:
- The type 2 Ins(1,4,5)P3 receptor (Ins(1,4,5)P3R) plays a crucial role in intracellular calcium signaling.
- Alternative splicing of mRNA transcripts generates diverse protein isoforms with potentially distinct functions.
Purpose of the Study:
- To identify and characterize novel isoforms of the type 2 Ins(1,4,5)P3R mRNA.
- To investigate the expression and potential function of a newly discovered isoform in muscle tissues.
Main Methods:
- Reverse transcriptase-mediated PCR (RT-PCR) was employed to detect novel mRNA isoforms.
- cDNA transfection into COS-7 cells and subsequent protein analysis using specific antibodies were performed.
- Western blot analysis was used to confirm protein expression and size.
Main Results:
- A novel exon inclusion led to the discovery of a unique type 2 Ins(1,4,5)P3R mRNA isoform.
- This isoform, expressed specifically in skeletal muscle and heart, encodes a truncated protein (TIPR) of 175 amino acids plus six novel residues.
- Transfected COS-7 cells expressed a 24 kDa TIPR protein recognized by a specific antibody; this isoform was also detected in human skeletal muscle and heart.
Conclusions:
- A novel muscle-specific Ins(1,4,5)P3R isoform (TIPR) has been identified in mice and humans.
- TIPR may play a regulatory role in the Ins(1,4,5)P3 signaling pathway within cardiac and skeletal muscles.