The repeat distance of myosin in the thick filaments of various muscles

The meridional spacing of the X-ray diffraction peak from the repeat of myosin along the thick filament of four muscles has been remeasured on the same apparatus. The frog sartorius gave a shorter repeat distance (143.7 A) than the three invertebrate muscles, which ranged from 144.9 to 145.4 A. These results confirm earlier measurements. Provided that the myosin molecules are staggered relative to one another by a constant 98 residues, it may be inferred that in vertebrate thick filaments part or all of the tail lies at a considerable angle to the filament axis, whereas in the invertebrates the angle is smaller.     

Tomographic 3D reconstruction of quick-frozen, Ca2+-activated contracting insect flight muscle

Motor actions of myosin were directly visualized by electron tomography of insect flight muscle quick-frozen during contraction. In 3D images, active cross-bridges are usually single myosin heads, bound preferentially to actin target zones sited midway between troponins. Active attached bridges (approximately 30% of all heads) depart markedly in axial and azimuthal angles from Rayment's rigor acto-S1 model, one-third requiring motor domain (MD) tilting on actin, and two-thirds keeping rigor contact with actin while the light chain domain (LCD) tilts axially from approximately 105 degrees to approximately 70 degrees. The results suggest the MD tilts and slews on actin from weak to strong binding, followed by swinging of the LCD through an approximately 35 degrees axial angle, giving an approximately 13 nm interaction distance and an approximately 4-6 nm working stroke.     

Purification and characterization of relaxin from the tammar wallaby (Macropus eugenii): bioactivity and expression in the corpus luteum

The objective of this study was to isolate and purify prorelaxin or mature relaxin from the tammar wallaby corpus luteum (CL), determine their structure and bioactivity, and test the hypothesis that enzymatic cleavage of prorelaxin occurs in late gestation. Tammar relaxin peptides were extracted from pooled corpora lutea of late pregnant tammars using a combination of HPLC methods, and they were identified using Western blotting with a human (H2) relaxin antisera and matrix-assisted laser desorption ionization time of flight mass spectrometry. Although no prorelaxin was identified, multiple 6-kDa peptides were detected, which corresponded to the predicted mature tammar relaxin amino acid sequence, with an A chain of 24 amino acids, and different B chain lengths of 28, 29, 30, and 32 amino acids. Tammar relaxin bound with high affinity to rat cortical relaxin receptors and stimulated cAMP production in the human monocytic cell line, THP-1, which expresses the relaxin receptor. Analysis of individual CL indicated that equivalent amounts of mature relaxin peptides were present throughout gestation and also in unmated tammars at equivalent stages of the luteal phase in the nonpregnant cycle. Immunoreactive relaxin was localized specifically to the luteal cells of the CL and the intensity of immunostaining did not vary between gestational stages. These data show that the CL of both pregnant and unmated tammar wallabies produces mature relaxin and suggests that relaxin expression in this species is not influenced by the conceptus. Moreover, the presence of mature relaxin throughout gestation implies that prohormone cleavage is not limited to the later stages of pregnancy     

Collagen studies in late pregnant relaxin null mice

The relaxin knockout (rlx -/-) mouse was used to assess the effect, during pregnancy, of relaxin with regard to water, collagen content, growth, and morphology of the nipple (N), vagina (V), uterus, cervix (C), pubic symphysis (PS), and mammary gland (MG). The results presented here indicate that during pregnancy, relaxin increases the growth of the N, C, V, and PS. Large increases in water content in the PS (20%) occurred in pregnant (Day 18.5) wild-type (rlx +/+) mice but not in rlx -/- animals. This indicates that in the PS, relaxin might increase the concentration of a water-retaining extracellular matrix component (hyaluronate). In the pregnant rlx +/+ mouse, collagen content decreased significantly in the N and V but not in other tissues. There were no significant changes in the rlx -/- mouse. This contrasts with findings in the rat, in which relaxin has been found to cause decreases in collagen concentrations in the V, C, and PS. Histological analysis showed that the collagen stain was more condensed in the tissues (V, C, PS, N, and MG) of rlx -/- mice than in those of rlx +/+ mice. This phenomenon indicates that the failure of collagen degradation and lack of growth in the N underlie the inability of the rlx -/- mice to feed their young, as reported previously. Vaginal and cervical luminal epithelia, which proliferated markedly in the rlx +/+ pregnant mice, remained relatively atrophic in the rlx -/- mice. As proliferation and differentiation of uterine and vaginal epithelia are thought to be induced by a paracrine stromal factor that acts upon estrogen stimulation, our results indicate that relaxin may be this paracrine factor.     

Evolution of the relaxin-like peptide family

Background  

The relaxin-like peptide family belongs in the insulin superfamily and consists of 7 peptides of high structural but low sequence similarity; relaxin-1, 2 and 3, and the insulin-like (INSL) peptides, INSL3, INSL4, INSL5 and INSL6. The functions of relaxin-3, INSL4, INSL5, INSL6 remain uncharacterised. The evolution of this family has been contentious; high sequence variability is seen between closely related species, while distantly related species show high similarity; an invertebrate relaxin sequence has been reported, while a relaxin gene has not been found in the avian and ruminant lineages.