Multiplicity of abnormal dystrophin in Becker muscular dystrophy: A Becker muscular dystrophy gene frequently produced two smaller sizes of dystrophin

Abstract

Dystrophin is a muscle cytoskeletal protein with a molecular mass (MM) of ∼ 420 kDa and an isoelectric point (p I) of ∼ 5.5, which is abnormal in size and/or abundance in Becker muscular dystrophy (BMD). We investigated the abnormality of dystrophin molecule in muscles biopsied from 23 BMD patients using the two-dimensional gel electrophoresis (TDGE). We found 7 protein spots which reacted specifically with the monoclonal anti-dystrophin antibody (mAb) A1C raised against N-terminal domain of the normal dystrophin. These spots were focused on the two-dimensional gel at the same position as the normal dystrophin (#1), at the position with MM ∼ 480 kDa pI ∼ 5.35 (#2), the position with MM ∼ 400−330 kDa pI ∼ 5.51−5.47 (#3), the position with MM ∼ 300 kDa pI ∼ 5.4 (#4), the position with MM ∼ 235−250 kDa pI ∼ 5.53−5.5 (#5), the position with MM ∼ 165 kDa pI ∼ 6.0 (#6), and the position with MM ∼ 160 kDa pI ∼ 5.75 (#7). These spots were classified into five patterns in individuals, that is, #1 alone in 3 patients, #3 alone in 1, the combination of # 3 and 5 in 17, the combination of # 1, 3 and 5 in 1 and the combination of # 1, 2, 4, 6 and 7 in 1. The combination of # 3 and 5 was observed in 17 of 23 patients (75%). In addition, both of # 3 and 5 reacted not only with the mAbs, A1C, Dys 3 and 5E2, which recognize the N-terminal domain of the normal dystrophin, but also with the mAbs, 4C5 and Dys 2, which recognize the C-terminal domain. Thus, each of # 3 and 5 preserved both N- and C-terminal domains of the normal dystrophin in spite of significant differences in MM and p I. Our observations conclude that the #5 is not a proteolytic fragment of the “full-length” dystrophin (#3), and suggest that some exons encoding triple helical segments at the central portion of dystrophin are spliced out to produce two abnormal dystrophins from a single mutated gene in the majority of BMD.