Biomedical subjects

J M Francois

Publications and source records attributed to J M Francois.

5 recordsLinked to original sources

PubMed1997

Horizontal glottectomy: functional and oncological results.

The authors report on their experience with 25 cases of horizontal glottectomy and discuss the functional and oncological results of this operation. Indications for this procedure are T1a and T1b glottic carcinomas. The overall three-year and five-year survival was respectively 94 and 88 per cent. Distant metastasis appears to be the major long-term failure, three patients dying of lung carcinoma. Short functional rehabilitation was observed in all cases. Decannulation was achieved in 100 per cent of the cases and none of our patients had laryngeal stenosis. The average time of removal of the nasogastric tube was 11.8 days. Because of its high local control and reduced functional after effects, horizontal glottectomy appears to be a reliable and safe procedure for limited glottic carcinomas and must be included in their therapeutic management.

Adult↗

PubMed1995

The functional role of the domains of troponin-C investigated with thrombin fragments of troponin-C reconstituted into skinned muscle fibers.

Proteolysis of rabbit fast skeletal troponin-C (RSTnC) with thrombin produces four separate fragments containing the following Ca2+-binding site(s): TH1 (residues 1-120) sites I-III; TH2 (121-159) site IV; TH3 (1-100) sites I and II; and TH4 (101-120) site III. We studied the ability of these fragments to restore the steady state isometric force in TnC-depleted skinned skeletal muscle fibers. Interestingly, we found that all investigated fragments of RSTnC possessed some of the properties of native RSTnC, but none of them could fully regulate contraction in the fibers like intact RSTnC. TH1 was the most effective in the force restoration (65%) whereas the smaller fragments developed about 50% (TH3 and TH4) or 20% (TH2) of the initial force of unextracted fibers. Additionally, much higher concentrations of TH2, TH3, and TH4 compared to RSTnC OR TH1 were necessary for force development suggesting a decreased affinity of these fragments to their binding site(s) in the fibers. Like intact RSTnC, TH1 was able to interact with the fibers in a Ca(2+)-independent (Mg(2+)-dependent) manner, indicating that at a minimum, Ca(2+)-binding site III is required for this type of binding. The initial binding of the other fragments to the TnC-depleted fibers occurred only in the presence of Ca2+. TH2 and TH4 appeared to bind to two different binding sites in the fibers. The binding to one of the sites caused partial force restoration. This binding of TH2 and TH4 was abolished when Ca2+ was removed. TH2 and TH4 binding to the second site required Ca2+ initially but was maintained in the presence of Mg2+. This interaction of TH2 and TH4 partially blocked the rebinding of RSTnC to the fibers. The latter results suggest that site III and IV in these small fragments, when removed from the constraints of the parent protein, may assume conformations that allow them to function, to a certain extent, like both the regulatory sites (I and II) and the Ca(2+)-Mg2+ sites(III and IV) of TnC.

Animals↗

PubMed1991

Effects of mutations in the central helix of troponin C on its biological activity.

To investigate the role of the central helix of skeletal muscle troponin C (TnC), five deletion mutants (Dobrowolski, Z., Xu, G.Q., and Hitchcock-DeGregori, S.E. (1991) J. Biol. Chem. 266, 5703-5710) of chicken TnC in the D/E linker region (K87EDAKGKSEEE97), dEDA, dKG, dKGK, dSEEE, and dKED-AKGK, were assayed for their ability to regulate muscle contraction by testing their effectiveness in restoring force and Ca2+ regulation to TnC-depleted rabbit skinned skeletal muscle fibers. By comparison with rabbit skeletal TnC, wild-type TnC, and chicken TnC, all mutants except dKG equally restored force development and Ca2+ regulation to TnC-depleted skinned muscle fibers. In contrast, approximately 4 times more dKG than rabbit skeletal TnC was required to reach 50% force restoration. Also, the pCa50 for dKG activation of force was significantly decreased. Thus, most of the TnC mutants that we studied did not have significantly altered biological activity in the skinned fiber assay. However, the 2-residue deletion in the central helix (dKG) significantly affected TnC activity. This deletion would be expected to produce a 160 degree rotation in the alpha-helix versus 60 degrees for dKGK and dEDA, 40 degrees in dSEEE, and 20 degrees in dKEDAKGK. Therefore, the change in orientation of the two Ca2(+)-binding domains appears to be a major parameter affecting TnC activity. The shift in the Ca2+ dependence in force activation may result from the inability of the Ca2(+)-specific domain to properly interact with its binding site on troponin I, an interaction which is known to increase the affinity of TnC for Ca2+ (Potter, J.D., and Gergely, J. (1975) J. Biol. Chem. 250, 4628-4633). In addition, the length of the central helix of TnC, Gly92, and the negatively charged cluster, EEE, appear not to be crucial for TnC activity.

Amino Acid Sequence↗

PubMed1991

Amino acid sequences and Ca2(+)-binding properties of two isoforms of barnacle troponin C.

Much of our knowledge of the relationship between elevation of free sarcoplasmic [Ca2+] and skeletal muscle contraction has come from physiological studies on barnacle muscle fibers. Little is known, however, about the biochemical properties of the barnacle proteins responsible for Ca2+ regulation. In order to help rectify this unfortunate situation, we purified the two major isoforms (BTnC1 and BTnC2) of troponin C (TnC) from the giant barnacle, Balanus nubilis, and determined their amino acid sequences. BTnC2, the more abundant isoform, contains 151 amino acid residues and has a calculated molecular weight of 16,838. Due to an elongated N-terminus, BTnC1 contains 158 amino acid residues and has a calculated molecular weight of 17,984. The two sequences can be aligned from their C-termini, with no insertions or deletions, but they are only 61% identical. Sequence differences are twice as frequent in the N-terminal halves as in the C-terminal halves but occur in all regions of the polypeptides. This indicates that BTnC1 and BTnC2 are products of different genes, rather than alternative transcripts of a single gene. Both isoforms contain the usual four ancestral Ca2(+)-binding regions, numbered I-IV from the N-terminus. Analysis of the sequences predicts that functional Ca2(+)-binding sites are present only in regions II and IV and that these sites are low-affinity Ca2(+)-specific type sites. Direct Ca2(+)-binding measurements using fluorescent Ca2+ indicators show that both isoforms bind 2 Ca2+/mol with equal affinity (KCa = 1.3 x 10(5) M-1) and the sites appear to be Ca2(+)-specific.

Amino Acid Sequence↗

PubMed1990

Evidence that both Ca(2+)-specific sites of skeletal muscle TnC are required for full activity.

To investigate the role of the Ca2(+)-specific (I and II) sites of fast skeletal muscle troponin C (TnC) in the regulation of contraction, we have produced two TnC mutants which have lost the ability to bind Ca2+ at either site I (VG1) or at site II (VG2). Both mutants were able to partially restore force to TnC-depleted skinned muscle fibers (approximately 25% for VG1 and approximately 50% for VG2). In contrast, bovine cardiac TnC (BCTnC), which like VG1 binds Ca2+ only at site II, could fully reactivate the contraction of TnC-depleted fibers. Higher concentrations of both mutants were required to restore force to the TnC-depleted fibers than with wild type TnC (WTnC) or BCTnC. VG1 and VG2 substituted fibers could not bind additional WTnC, indicating that all of the TnC-binding sites were saturated with the mutant TnC's. The Ca2+ concentration required for force activation was much higher for VG1 and VG2 substituted fibers than for WTnC or BCTnC substituted fibers. Also, the steepness of force activation was much less in VG1 and VG2 versus WTnC and BCTnC substituted fibers. These results suggest cooperative interactions between sites I and II in WTnC. In contrast, BCTnC has essentially the same apparent Ca2+ affinity and steepness of force activation as does WTnC. Thus, cardiac TnC must have structural differences from WTnC which compensate for the lack of site I, while in WTnC, both Ca2(+)-specific sites are probably crucial for full functional activity.

Amino Acid Sequence↗