Contractile properties and proteins of smooth muscles of a calponin knockout mouse

Abstract

1. The role of h1-calponin in regulating the contractile properties of smooth muscle was investigated in bladder and vas deferens of mice carrying a targeted mutation in both alleles designed to inactivate the basic calponin gene. These calponin knockout (KO) mice displayed no detectable h1-calponin in their smooth muscles.2. The amplitudes of Ca 2+ sensitization, force and Ca 2+ sensitivity were not significantly different in permeabilized smooth muscle of KO compared with wild-type (WT) mice, nor were the delays in onset and half-times of Ca 2+ sensitization, initiated by flash photolysis of caged GTPyS, different.3. The unloaded shortening velocity (V us ) of thiophosphorylated fibres was significantly (P < 0.05) faster in the smooth muscle of KO than WT animals, but could be slowed by exogenous calponin to approximate WT levels; the concentration dependence of exogenous calponin slowing of V us was proportional to its actomyosin binding in situ.4. Actin expression was reduced by 25-50%, relative to that of myosin heavy chain, in smooth muscle of KO mice, without any change in the relative distribution of the actin isoforms.5. We conclude that the faster V us of smooth muscle of the KO mouse is consistent with, but does not prove without further study, physiological regulation of the crossbridge cycle by calponin. Our results show no detectable role of calponin in the signal transduction of the Ca 2+ -sensitization pathways in smooth muscle.

Myoepithelial Cell Differentiation Markers in Ductal Carcinoma in Situ Progression

“…9,10,36e39 However, the question of whether progressive loss of myoepithelial cell differentiation markers occurs before invasive disease has not been thoroughly investigated. Here, we used IHC to assess a panel of myoepithelial cell differentiation markers, a-SMA, the major smooth muscle contractile protein; calponin, an actin-binding protein that regulates the power stroke during smooth muscle contraction 40,41 ; and p63, a transcription factor with putative tumor suppressor function. 10,12,42 In nonetumor-bearing murine ducts, as expected, evidence was found for uniform myoepithelial cell expression of a-SMA ( Figure 4A), calponin ( Figure 4B), and p63 ( Figure 4C).…”

“…However, gain of a-SMA staining in adjacent cancerassociated fibroblasts limits the usefulness of a-SMA in the diagnosis of DCIS. 48,49 Calponin, an important actin-myosin regulator in smooth muscle cells, 40,41 shows more frequent loss within myoepithelial cells, being largely present in lesions 4 weeks after injection and lost by 10 weeks after injection. Our data suggest that myoepithelial cell loss of calponin occurs before loss of a-SMA, but after loss of p63, which occurs by 4 weeks after tumor cell injection in our murine model.…”

“…Calponin is an integral component of a-SMA where it regulates myosin binding to actin and modulates the power stroke during smooth muscle contraction. 40,41 In addition, calponin has an identified signaling function, because calponin knockout results in 25% to 50% loss in actin gene expression in smooth muscle cells. 40 Because both of these calponin-dependent functions are anticipated to contribute to structural integrity of the myoepithelium, calponin down-regulation is consistent with compromised myoepithelium and possible tumor cell escape.…”

“…40,41 In addition, calponin has an identified signaling function, because calponin knockout results in 25% to 50% loss in actin gene expression in smooth muscle cells. 40 Because both of these calponin-dependent functions are anticipated to contribute to structural integrity of the myoepithelium, calponin down-regulation is consistent with compromised myoepithelium and possible tumor cell escape. However, these identified smooth muscle functions of calponin do not readily explain the gain in p63 we observed in intraductal tumor cells adjacent to calponin-negative myoepithelial cells.…”

Altered Mechanical Properties in Smooth Muscle of Mice with a Mutated Calponin Locus

“…Some lines of evidence have been accumulated to indicate the in vitro biochemical properties of calponin (Haeberle et al ., 1994;Hodgkinson et al ., 1997;Malmqvist et al ., 1997). To elucidate the function of calponin physiologically, mice with a mutated basic calponin locus were generated and the mechanical properties of smooth muscle in intact tissue were compared between calponin knockout (KO) mice and wild type (WT) one Matthew et al ., 2000;Takahashi et al ., 2000). The present experiment is a further detailed study of the mechanical properties, especially about force development in the smooth muscles of aorta and vas deferens in intact tissues in calponin KO and WT mice.…”