Calpain-2 specifically cleaves Junctophilin-2 at the same site as Calpain-1 but with less efficacy. Issue 19 (15th October 2021)

• Record Type: Journal Article
• Authors: Wang, Jinxi; Ciampa, Grace; Zheng, Dong; Shi, Qian; Chen, Biyi; Abel, E. Dale; Peng, Tianqing; Hall, Duane D.; Song, Long-Sheng
• Is Part Of: Biochemical journal. Volume 478:Issue 19(2021)
• Page Start: 3539
• Page End: 3553
• Publication Date: 2021-10-15
• Subjects: Calpain-1 -- Calpain-2 -- heart failure -- Junctophilin-2; Biochemistry -- Periodicals; 572
• DOI: 10.1042/BCJ20210629
• Languages: English
• ISSNs: 0264-6021

Abstract

Abstract : Calpain proteolysis contributes to the pathogenesis of heart failure but the calpain isoforms responsible and their substrate specificities have not been rigorously defined. One substrate, Junctophilin-2 (JP2), is essential for maintaining junctional cardiac dyads and excitation-contraction coupling. We previously demonstrated that mouse JP2 is cleaved by calpain-1 (CAPN1) between Arginine 565 (R 565 ) and Threonine 566 (T 566 ). Recently, calpain-2 (CAPN2) was reported to cleave JP2 at a novel site between Glycine 482 (G 482 ) and Threonine 483 (T 483 ). We aimed to directly compare the contributions of each calpain isoform, their Ca 2+ sensitivity, and their cleavage site selection for JP2. We find CAPN1, CAPN2 and their requisite CAPNS1 regulatory subunit are induced by pressure overload stress that is concurrent with JP2 cleavage. Using in vitro calpain cleavage assays, we demonstrate that CAPN1 and CAPN2 cleave JP2 into similar 75 kD N-terminal (JP2NT) and 25 kD C-terminal fragments (JP2CT) with CAPNS1 co-expression enhancing proteolysis. Deletion mutagenesis shows both CAPN1 and CAPN2 require R 565 /T 566 but not G 482 /T 483 . When heterologously expressed, the JP2CT peptide corresponding to R 565 /T 566 cleavage approximates the 25 kD species found during cardiac stress while the C-terminal peptide from potential cleavage at G 482 /T 483 produces a 35 kD product. Similar results were obtained for human JP2. Finally, we show that CAPN1 has higher Ca 2+ sensitivity and cleavage efficacy than CAPN2 on JP2 and other cardiac substrates including cTnT, cTnI and β2-spectrin. We conclude that CAPN2 cleaves JP2 at the same functionally conserved R 565 /T 566 site as CAPN1 but with less efficacy and suggest heart failure may be targeted through specific inhibition of CAPN1.