The myosin interacting-heads motif is present in the relaxed thick filament of the striated muscle of scorpion.

Electron microscopy (EM) studies of 2D crystals of smooth muscle myosin molecules have shown that in the inactive state the two heads of a myosin molecule interact asymmetrically forming a myosin interacting-heads motif. This suggested that inactivation of the two heads occurs by blocking of the actin-binding site of one (free head) and the ATP hydrolysis site of the other (blocked head). This motif has been found by EM of isolated negatively stained myosin molecules of unregulated (vertebrate skeletal and cardiac muscle) and regulated (invertebrate striated and vertebrate smooth muscle) myosins, and nonmuscle myosin. The same motif has also been found in 3D-reconstructions of frozen-hydrated (tarantula, Limulus, scallop) and negatively stained (scallop, vertebrate cardiac) isolated thick filaments. We are carrying out studies of isolated thick filaments from other species to assess how general this myosin interacting-heads motif is. Here, using EM, we have visualized isolated, negatively stained thick filaments from scorpion striated muscle. We modified the iterative helical real space reconstruction (IHRSR) method to include filament tilt, and band-pass filtered the aligned segments before averaging, achieving a 3.3 nm resolution 3D-reconstruction. This reconstruction revealed the presence of the myosin interacting-heads motif (adding to evidence that is widely spread), together with 12 subfilaments in the filament backbone. This demonstrates that conventional negative staining and imaging can be used to detect the presence of the myosin interacting-heads motif in helically ordered thick filaments from different species and muscle types, thus avoiding the use of less accessible cryo-EM and low electron-dose procedures.

Differences in the nuclear chromatin among various stages of the life cycle of Trypanosoma cruzi.

Trypanosoma cruzi is the etiological agent of Chagas. Although the nuclear chromatin of this parasite is organized in the form of nucleosome filaments, its chromatin is physically and enzymatically fragile, and no condensation into chromosomes occurs during mitosis. All previous investigations have been carried out with epimastigote form in its proliferate stage. It is not known whether these differences in chromatin structure are also found in the non-proliferate stationary epimastigote forms and in tissue derived trypomastigotes. Our results confirm that chromatin of logarithmic epimastigotes presents limited compaction when increasing salt concentrations from 1 to 100 mM NaCl, and no 30-nm fibers were formed. Contrary to these results, non-proliferative forms of the parasites showed a pattern of compactation similar to that observed in rat liver chromatin, where solenoids of 30-nm fibers are formed at 100-mM NaCl. In accordance with these results, digestion of the nuclear chromatin with DNase I revealed that the chromatin of logarithmic phase epimastigotes was more accessible to the enzyme. We conclude from these results that structural differences in the chromatin exist not only between T. cruzi and higher eukaryotes but also among various forms of the parasite. The functional significance of these differences are currently under investigation.

Una infección viral en la glándula cervical de larvas de Dione junio (Lepidoptera: Nymphalidae) / A viral infection in the neck glandula of larvae of Dione junio (Lepidoptera: Nymphalidae)

Lepidopteran larvae may be attacked by different viruses, many of which belong to the Baculoviridae family. Whilst studying the ultrastructure of the neck gland in Dione junio larvae we found that in later instars the larvae showed symptoms of attack by two types of virus. The glands were prepared for optical and electron microscopy using sodium cacodylate buffer and standard procedures (0.1M, pH 7.2). The neck gland is composed of two oval internal sacks which communicate with the exterior via an extracellular channel. Each sack contains, in its external region, cells with large, irregular nuclei and a dense cytoplasm containing numerous small mitochondria. In infected larvae, the tissues are damaged and the nuclear polyhedrosis virus can be observed in several of the nuclei. In the cytoplasm another "rickettsia type" virus, may be observed. The pathogenic viruses present in D. junio larvae could be studied as potential biological controls of this pest.