The ability to coordinate the timing of motor protein activation lies at the center of a wide range of cellular motile processes including endocytosis, cell division, and cancer cell migration. We show that calcium dramatically alters the conformation and activity of the myosin-VI motor implicated in pivotal steps of these processes. We resolved the change in motor conformation and in structural flexibility using single particle analysis of electron microscopic data and identified interacting domains using fluorescence spectroscopy. We discovered that calcium binding to calmodulin increases the binding affinity by a factor of 2,500 for a bipartite binding site on myosin-VI. The ability of calcium-calmodulin to seek out and bridge between binding site components directs a major rearrangement of the motor from a compact dormant state into a cargo binding primed state that is nonmotile. The lack of motility at high calcium is due to calmodulin switching to a higher affinity binding site, which leaves the original IQ-motif exposed, thereby destabilizing the lever arm. The return to low calcium can either restabilize the lever arm, required for translocating the cargobound motors toward the center of the cell, or refold the cargo-free motors into an inactive state ready for the next cellular calcium flux.unconventional myosin | electron microscopy | calmodulin I n human cells, cytoskeletal motor proteins move along microtubules and actin filaments to generate complex cellular functions that require a precise timing of motor activation and inactivation. Myosin-VI is thought to have unique properties because it is the only myosin in the human genome shown to move toward the minus end of actin filaments (1). Apart from its roles in the formation of stereocilia in cells of the auditory system (2, 3), membrane internalization (4-6), and delivery of membrane to the leading edge in migratory cells (7), myosin-VI is an early marker of cancer development, aggressiveness, and cancer-cell invasion because of its dramatically up-regulated expression in breast, lung, prostate, ovary, and gastresophagus carcinoma cells (7-11). How this motor might promote cancercell migration, proliferation, and survival is unknown.In migrating cells, localized calcium transients (∼50 nM to ∼10 μM) (12, 13) have been reported to play a multifunctional role in steering directional movement (14), cytoskeleton redistribution, and relocation of focal adhesions (15). The effect of calcium transients on the mobilization and cargo binding of myosin-VI and on its mechanical activation, however, are not understood. In the current model, the catalytic head domain hydrolyzes ATP, whereas the tail domain anchors the motor to specific compartments. In vitro studies have shown that calcium affects myosin-VI binding to phospholipids (6), as well as the kinetics and motility rate of the motor (16, 17). The underlying molecular mechanisms, however, are unknown. It has also been discussed that myosin-VI might be able to adopt an inactive folded state (18,19), perha...

Myosin XXI is the only myosin expressed in Leishmania parasites. Although it is assumed that it performs a variety of motile functions, the motor's oligomerization states, cargo-binding, and motility are unknown. Here we show that binding of a single calmodulin causes the motor to adopt a monomeric state and to move actin filaments. In the absence of calmodulin, nonmotile dimers that cross-linked actin filaments were formed. Unexpectedly, structural analysis revealed that the dimerization domains include the calmodulin-binding neck region, essential for the generation of force and movement in myosins. Furthermore, monomeric myosin XXI bound to mixed liposomes, whereas the dimers did not. Lipid-binding sections overlapped with the dimerization domains, but also included a phox-homology domain in the converter region. We propose a mechanism of myosin regulation where dimerization, motility, and lipid binding are regulated by calmodulin. Although myosin-XXI dimers might act as nonmotile actin cross-linkers, the calmodulin-binding monomers might transport lipid cargo in the parasite.unconventional myosin | motor properties

Sex determination of birds is important for many ecological studies but is often difficult in species with monomorphic plumage. Morphology often provides a possibility for sex determination, but the characters need to be verified. We tested whether five passerine species can be sexed according to standard morphological measurements applying a forward logistic regression with sex determined by molecular analysis as the dependent variable. Furthermore, we tested whether the results can be used on a larger geographic scale by applying morphological sexing methods gained by similar studies from other regions to our data set. Of the five species of this study only Garden Warblers Sylvia borin could not be sexed morphologically. In the Robin Erithacus rubecula, 87.2% of all individuals were sexed correctly. For Reed Warblers Acrocephalus scirpaceus, Willow Warblers Phylloscopus trochilus and Reed Buntings Emberiza schoeniclus, the respective values were 77.6, 89.4 and 86.4%. When the logistic regression functions from similar studies on Robins and Reed Buntings in Denmark and Scotland were applied to the birds from south-western Germany, they performed less well compared to the original dataset of these studies and compared to the logistic regression function of our own study. The same was the case for Willow Warblers when a wing length criterion used in Great Britain was applied to the birds of our study. These discrepancies may have several explanations: (1) the models are optimised for the dataset from which they were extracted, (2) inter-ringer variation in measurements, (3) the use of different age cohorts, (4) different morphology due to different habitat availability around the study site, or, most likely, (5) different morphology due to different migratory behaviour. We recommend that morphological sex differentiation methods similar to this study (1) be only used population specific, (2) only with one age cohort and (3) to adjust the extracted equations from time to time.

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