Endothelin receptor Aa regulates proliferation and differentiation of Erb-dependent pigment progenitors in zebrafish.

Skin pigment patterns are important, being under strong selection for multiple roles including camouflage and UV protection. Pigment cells underlying these patterns form from adult pigment stem cells (APSCs). In zebrafish, APSCs derive from embryonic neural crest cells, but sit dormant until activated to produce pigment cells during metamorphosis. The APSCs are set-aside in an ErbB signaling dependent manner, but the mechanism maintaining quiescence until metamorphosis remains unknown. Mutants for a pigment pattern gene, parade, exhibit ectopic pigment cells localised to the ventral trunk, but also supernumerary cells restricted to the Ventral Stripe. Contrary to expectations, these melanocytes and iridophores are discrete cells, but closely apposed. We show that parade encodes Endothelin receptor Aa, expressed in the blood vessels, most prominently in the medial blood vessels, consistent with the ventral trunk phenotype. We provide evidence that neuronal fates are not affected in parade mutants, arguing against transdifferentiation of sympathetic neurons to pigment cells. We show that inhibition of BMP signaling prevents specification of sympathetic neurons, indicating conservation of this molecular mechanism with chick and mouse. However, inhibition of sympathetic neuron differentiation does not enhance the parade phenotype. Instead, we pinpoint ventral trunk-restricted proliferation of neural crest cells as an early feature of the parade phenotype. Importantly, using a chemical genetic screen for rescue of the ectopic pigment cell phenotype of parade mutants (whilst leaving the embryonic pattern untouched), we identify ErbB inhibitors as a key hit. The time-window of sensitivity to these inhibitors mirrors precisely the window defined previously as crucial for the setting aside of APSCs in the embryo, strongly implicating adult pigment stem cells as the source of the ectopic pigment cells. We propose that a novel population of APSCs exists in association with medial blood vessels, and that their quiescence is dependent upon Endothelin-dependent factors expressed by the blood vessels.

Zebrafish Bioassay for Screening Therapeutic Candidates Based on Melanotrophic Activity.

In this study, we used the zebrafish animal model to establish a bioassay by which physiological efficacy differential of alpha-melanocyte-stimulating hormone (α-MSH) analogues could be measured by melanosome dispersion in zebrafish larvae. Brain-skin connection research has purported the interconnectedness between the nervous system and skin physiology. Accordingly, the neuropeptide α-MSH is a key regulator in several physiological processes, such as skin pigmentation in fish. In mammals, α-MSH has been found to regulate motivated behavior, appetite, and emotion, including stimulation of satiety and anxiety. Several clinical and animal model studies of autism spectrum disorder (ASD) have already demonstrated the effectiveness of α-MSH in restoring the social deficits of autism. Therefore, we sought to analyze the effect of synthetic and naturally-occurring α-MSH variants amongst different species. Our results showed that unique α-MSH derivatives from several fish species produced differential effects on the degree of melanophore dispersion. Using α-MSH human form as a standard, we could identify derivatives that induced greater physiological effects; particularly, the synthetic analogue melanotan-II (MT-II) exhibited a higher capacity for melanophore dispersion than human α-MSH. This was consistent with previous findings in an ASD mouse model demonstrating the effectiveness of MT-II in improving ASD behavioral symptoms. Thus, the melanophore assay may serve as a useful screening tool for therapeutic candidates for novel drug discovery.

Melanophore multinucleation pathways in zebrafish.

In zebrafish, apart from mononuclear melanophores, bi- and trinuclear melanophores are frequently observed; however, the manner in which multinucleation of these cells occurs during fish development remains unknown. Here, we analyzed the processes underlying multinucleation of zebrafish melanophores. Transgenic zebrafish in which melanophore nuclei were labeled with a histone H2B-red fluorescent reporter protein were used to evaluate the distribution of mono-, bi-, and trinuclear melanophores in both the trunk and fin. Half of the melanophores examined were binuclear and approximately 1% were trinuclear. We compared cell size, cell motility, and survival rate between mono- and binuclear melanophores grown in a culture dish, and we found that cell size and survival rate were significantly larger in binuclear melanophores. We then analyzed the behavior of melanoblasts and melanophores from transgenic zebrafish using in vivo and in vitro live-cell imaging. We detected division and differentiation of melanoblasts, as well as melanoblast nuclear division without subsequent cellular division. In addition, we observed cellular and nuclear division in melanophores, although these events were very infrequent in vitro. On the basis of our findings, we present a scheme for melanophore multinucleation in zebrafish.

Insights from zebrafish on human pigment cell disease and treatment.

Black pigment cells, melanocytes, arise early during development from multipotent neural crest cells. Melanocytes protect human skin from DNA damaging sunrays and provide color for hair, eyes, and skin. Several disorders and diseases originate from these cells, including the deadliest skin cell cancer, melanoma. Thus, melanocytes are critical for a healthy life and for protecting humans from disease. Due to the ease of visualizing pigment cells through transparent larvae skin and conserved roles for zebrafish melanophore genes to mammalian melanocyte genes, zebrafish larvae offer a biologically relevant model for understanding pigment cell development and disease in humans. This review discusses our current knowledge of melanophore biology and how zebrafish are contributing to improving how diseases of melanocytes are understood and treated in humans. Developmental Dynamics 246:889-896, 2017. © 2017 Wiley Periodicals, Inc.

Involvement of Delta/Notch signaling in zebrafish adult pigment stripe patterning.

The skin pigment pattern of zebrafish is a good model system in which to study the mechanism of biological pattern formation. Although it is known that interactions between melanophores and xanthophores play a key role in the formation of adult pigment stripes, molecular mechanisms for these interactions remain largely unknown. Here, we show that Delta/Notch signaling contributes to these interactions. Ablation of xanthophores in yellow stripes induced the death of melanophores in black stripes, suggesting that melanophores require a survival signal from distant xanthophores. We found that deltaC and notch1a were expressed by xanthophores and melanophores, respectively. Moreover, inhibition of Delta/Notch signaling killed melanophores, whereas activation of Delta/Notch signaling ectopically in melanophores rescued the survival of these cells, both in the context of pharmacological inhibition of Delta/Notch signaling and after ablation of xanthophores. Finally, we showed by in vivo imaging of cell membranes that melanophores extend long projections towards xanthophores in the yellow stripes. These data suggest that Delta/Notch signaling is responsible for a survival signal provided by xanthophores to melanophores. As cellular projections can enable long-range interaction between membrane-bound ligands and their receptors, we propose that such projections, combined with direct cell-cell contacts, can substitute for the effect of a diffusible factor that would be expected by the conventional reaction-diffusion (Turing) model.

In vitro analysis suggests that difference in cell movement during direct interaction can generate various pigment patterns in vivo.

Pigment patterns of organisms have invoked strong interest from not only biologists but also, scientists in many other fields. Zebrafish is a useful model animal for studying the mechanism of pigment pattern formation. The zebrafish stripe pattern is primarily two types of pigment cells: melanophores and xanthophores. Previous studies have reported that interactions among these pigment cells are important for pattern formation. In the recent report, we found that the direct contact by xanthophores induces the membrane depolarization of melanophores. From analysis of jaguar mutants, it is suggested that the depolarization affects the movements of melanophores. To analyze the cell movement in detail, we established a unique in vitro system. It allowed us to find that WT xanthophores induced repulsive movement of melanophores through direct contact. The xanthophores also chased the melanophores. As a result, they showed run-and-chase movements. We also analyzed the cell movement of pigment cells from jaguar and leopard mutants, which have fuzzy stripes and spot patterns, respectively. jaguar cells showed inhibited run-and-chase movements, and leopard melanophores scarcely showed repulsive response. Furthermore, we paired mutant and WT cells and showed which of the melanophores and xanthophores have responsibility for the altered cell movements. These results suggested that there is a correspondence relationship between the cell movements and pigment patterns. The correspondence relationship highlighted the importance of the cell movements in the pattern formation and showed that our system is a quite useful system for future study in this field.

[Regeneration of the Skin Pigment System during Larval Development of the Clawed Frog].

We demonstrate regeneration capability of the skin pigment system of clawed frog larvae after local damage to melanophores without skin rupture. The contribution to recovery of pigmentation of the injured area of de novo differentiation of melanophores is compared to contribution of mitotic division of undamaged melanophores localized on the boundaries of the injured area. The regeneration process is observed during various stages of pigment system development of larvae. We establish that, compared to ontogenetic dynamics, pigmentation development in animals is more intense during the regeneration.

Self-organized optical device driven by motor proteins.

Protein molecules produce diverse functions according to their combination and arrangement as is evident in a living cell. Therefore, they have a great potential for application in future devices. However, it is currently very difficult to construct systems in which a large number of different protein molecules work cooperatively. As an approach to this challenge, we arranged protein molecules in artificial microstructures and assembled an optical device inspired by a molecular system of a fish melanophore. We prepared arrays of cell-like microchambers, each of which contained a scaffold of microtubule seeds at the center. By polymerizing tubulin from the fixed microtubule seeds, we obtained radially arranged microtubules in the chambers. We subsequently prepared pigment granules associated with dynein motors and attached them to the radial microtubule arrays, which made a melanophore-like system. When ATP was added to the system, the color patterns of the chamber successfully changed, due to active transportation of pigments. Furthermore, as an application of the system, image formation on the array of the optical units was performed. This study demonstrates that a properly designed microstructure facilitates arrangement and self-organization of molecules and enables assembly of functional molecular systems.

Interactions with iridophores and the tissue environment required for patterning melanophores and xanthophores during zebrafish adult pigment stripe formation.

Skin pigment patterns of vertebrates are a classic system for understanding fundamental mechanisms of morphogenesis, differentiation, and pattern formation, and recent studies of zebrafish have started to elucidate the cellular interactions and molecular mechanisms underlying these processes. In this species, horizontal dark stripes of melanophores alternate with light interstripes of yellow or orange xanthophores and iridescent iridophores. We showed previously that the highly conserved zinc finger protein Basonuclin-2 (Bnc2) is required in the environment in which pigment cells reside to promote the development and maintenance of all three classes of pigment cells; bnc2 mutants lack body stripes and interstripes. Previous studies also revealed that interactions between melanophores and xanthophores are necessary for organizing stripes and interstripes. Here we show that bnc2 promotes melanophore and xanthophore development by regulating expression of the growth factors Kit ligand a (Kitlga) and Colony stimulating factor-1 (Csf1), respectively. Yet, we found that rescue of melanophores and xanthophores was insufficient for the recovery of stripes in the bnc2 mutant. We therefore asked whether bnc2-dependent iridophores might contribute to stripe and interstripe patterning as well. We found that iridophores themselves express Csf1, and by ablating iridophores in wild-type and mutant backgrounds, we showed that iridophores contribute to organizing both melanophores and xanthophores during the development of stripes and interstripes. Our results reveal an important role for the cellular environment in promoting adult pigment pattern formation and identify new components of a pigment-cell autonomous pattern-generating system likely to have broad implications for understanding how pigment patterns develop and evolve.

Ferritin H subunit gene is specifically expressed in melanophore precursor-derived white pigment cells in which reflecting platelets are formed from stage II melanosomes in the periodic albino mutant of Xenopus laevis.

"White pigment cells" are derived from melanophore precursors and contain both melanophore-specific and iridophore-specific pigment organelles. Whereas melanophores differentiate in the wild type regenerating tail, white pigment cells appear in the regenerating tail in the periodic albino mutant (a(p)/a(p)) of Xenopus laevis. The localization and density of white pigment cells in the mutant regenerating tail are similar to those of melanophores in the wild type regenerating tail. Here, white pigment cells in the mutant regenerating tail have been compared with melanophores in the wild type regenerating tail in the presence of phenylthiourea (PTU), which inhibits melanosome maturation in melanophores but does not affect reflecting platelet formation in white pigment cells. Ultrastructural analysis shows that reflecting platelet formation in white pigment cells is different from that in iridophores. Reflecting platelets in iridophores are formed from spherical vesicles with electron-dense material, whereas they are formed from stage II melanosomes characteristic of melanophore precursors in white pigment cells. Ultrastructural features of pigment organelles, except reflecting platelets, are similar between mutant melanophores and white pigment cells. In an attempt to identify specific genes in white pigment cells, a subtracted cDNA library enriched for mutant cDNAs has been prepared. Subtracted cDNA fragments have been cloned and selected by whole mount in situ hybridization. Among cDNA fragments examined so far, the ferritin H subunit gene is specifically expressed in white pigment cells, but not in melanophores. Pigment organellogenesis and specific gene expression in white pigment cells are also discussed.

Comparison of pigment cell ultrastructure and organisation in the dermis of marble trout and brown trout, and first description of erythrophore ultrastructure in salmonids.

Skin pigmentation in animals is an important trait with many functions. The present study focused on two closely related salmonid species, marble trout (Salmo marmoratus) and brown trout (S. trutta), which display an uncommon labyrinthine (marble-like) and spot skin pattern, respectively. To determine the role of chromatophore type in the different formation of skin pigment patterns in the two species, the distribution and ultrastructure of chromatophores was examined with light microscopy and transmission electron microscopy. The presence of three types of chromatophores in trout skin was confirmed: melanophores; xanthophores; and iridophores. In addition, using correlative microscopy, erythrophore ultrastructure in salmonids was described for the first time. Two types of erythrophores are distinguished, both located exclusively in the skin of brown trout: type 1 in black spot skin sections similar to xanthophores; and type 2 with a unique ultrastructure, located only in red spot skin sections. Morphologically, the difference between the light and dark pigmentation of trout skin depends primarily on the position and density of melanophores, in the dark region covering other chromatophores, and in the light region with the iridophores and xanthophores usually exposed. With larger amounts of melanophores, absence of xanthophores and presence of erythrophores type 1 and type L iridophores in the black spot compared with the light regions and the presence of erythrophores type 2 in the red spot, a higher level of pigment cell organisation in the skin of brown trout compared with that of marble trout was demonstrated. Even though the skin regions with chromatophores were well defined, not all the chromatophores were in direct contact, either homophilically or heterophilically, with each other. In addition to short-range interactions, an important role of the cellular environment and long-range interactions between chromatophores in promoting adult pigment pattern formation of trout are proposed.

Melanophore migration and survival during zebrafish adult pigment stripe development require the immunoglobulin superfamily adhesion molecule Igsf11.

The zebrafish adult pigment pattern has emerged as a useful model for understanding the development and evolution of adult form as well as pattern-forming mechanisms more generally. In this species, a series of horizontal melanophore stripes arises during the larval-to-adult transformation, but the genetic and cellular bases for stripe formation remain largely unknown. Here, we show that the seurat mutant phenotype, consisting of an irregular spotted pattern, arises from lesions in the gene encoding Immunoglobulin superfamily member 11 (Igsf11). We find that Igsf11 is expressed by melanophores and their precursors, and we demonstrate by cell transplantation and genetic rescue that igsf11 functions autonomously to this lineage in promoting adult stripe development. Further analyses of cell behaviors in vitro, in vivo, and in explant cultures ex vivo demonstrate that Igsf11 mediates adhesive interactions and that mutants for igsf11 exhibit defects in both the migration and survival of melanophores and their precursors. These findings identify the first in vivo requirements for igsf11 as well as the first instance of an immunoglobulin superfamily member functioning in pigment cell development and patterning. Our results provide new insights into adult pigment pattern morphogenesis and how cellular interactions mediate pattern formation.

Lateral motion and bending of microtubules studied with a new single-filament tracking routine in living cells.

The cytoskeleton is involved in numerous cellular processes such as migration, division, and contraction and provides the tracks for transport driven by molecular motors. Therefore, it is very important to quantify the mechanical behavior of the cytoskeletal filaments to get a better insight into cell mechanics and organization. It has been demonstrated that relevant mechanical properties of microtubules can be extracted from the analysis of their motion and shape fluctuations. However, tracking individual filaments in living cells is extremely complex due, for example, to the high and heterogeneous background. We introduce a believed new tracking algorithm that allows recovering the coordinates of fluorescent microtubules with ∼9 nm precision in in vitro conditions. To illustrate potential applications of this algorithm, we studied the curvature distributions of fluorescent microtubules in living cells. By performing a Fourier analysis of the microtubule shapes, we found that the curvatures followed a thermal-like distribution as previously reported with an effective persistence length of ∼20 µm, a value significantly smaller than that measured in vitro. We also verified that the microtubule-associated protein XTP or the depolymerization of the actin network do not affect this value; however, the disruption of intermediate filaments decreased the persistence length. Also, we recovered trajectories of microtubule segments in actin or intermediate filament-depleted cells, and observed a significant increase of their motion with respect to untreated cells showing that these filaments contribute to the overall organization of the microtubule network. Moreover, the analysis of trajectories of microtubule segments in untreated cells showed that these filaments presented a slower but more directional motion in the cortex with respect to the perinuclear region, and suggests that the tracking routine would allow mapping the microtubule dynamical organization in cells.

Endothelin modulates the circadian expression of non-visual opsins.

The non-visual opsin, melanopsin, expressed in the mammalian retina, is considered a circadian photopigment because it is responsible to entrain the endogenous biological clock. This photopigment is also present in the melanophores of Xenopus laevis, where it was first described, but its role in these cells is not fully understood. X. laevis melanophores respond to light with melanin granule dispersion, the maximal response being achieved at the wavelength of melanopsin maximal excitation. Pigment dispersion can also be triggered by endothelin-3 (ET-3). Here we show that melanin translocation is greater when a blue light pulse was applied in the presence of ET-3. In addition, we demonstrated that mRNA levels of the melanopsins Opn4x and Opn4m exhibit temporal variation in melanophores under light/dark (LD) cycles or constant darkness, suggesting that this variation is clock-driven. Moreover, under LD cycles the oscillations of both melanopsins show a circadian profile suggesting a role for these opsins in the photoentrainment mechanism. Blue-light pulse decreased Opn4x expression, but had no effect on Opn4m. ET-3 abolishes the circadian rhythm of expression of both opsins; in addition the hormone increases Opn4x expression in a dose-, circadian time- and light-dependent way. ET-3 also increases the expression of its own receptor, in a dose-dependent manner. The variation of melanopsin levels may represent an adaptive mechanism to ensure greater melanophore sensitivity in response to environmental light conditions with ideal magnitude in terms of melanin granule dispersion, and consequently color change.

Post-embryonic nerve-associated precursors to adult pigment cells: genetic requirements and dynamics of morphogenesis and differentiation.

The pigment cells of vertebrates serve a variety of functions and generate a stunning variety of patterns. These cells are also implicated in human pathologies including melanoma. Whereas the events of pigment cell development have been studied extensively in the embryo, much less is known about morphogenesis and differentiation of these cells during post-embryonic stages. Previous studies of zebrafish revealed genetically distinct populations of embryonic and adult melanophores, the ectotherm homologue of amniote melanocytes. Here, we use molecular markers, vital labeling, time-lapse imaging, mutational analyses, and transgenesis to identify peripheral nerves as a niche for precursors to adult melanophores that subsequently migrate to the skin to form the adult pigment pattern. We further identify genetic requirements for establishing, maintaining, and recruiting precursors to the adult melanophore lineage and demonstrate novel compensatory behaviors during pattern regulation in mutant backgrounds. Finally, we show that distinct populations of latent precursors having differential regenerative capabilities persist into the adult. These findings provide a foundation for future studies of post-embryonic pigment cell precursors in development, evolution, and neoplasia.

Differentiation of zebrafish melanophores depends on transcription factors AP2 alpha and AP2 epsilon.

A model of the gene-regulatory-network (GRN), governing growth, survival, and differentiation of melanocytes, has emerged from studies of mouse coat color mutants and melanoma cell lines. In this model, Transcription Factor Activator Protein 2 alpha (TFAP2A) contributes to melanocyte development by activating expression of the gene encoding the receptor tyrosine kinase Kit. Next, ligand-bound Kit stimulates a pathway activating transcription factor Microphthalmia (Mitf), which promotes differentiation and survival of melanocytes by activating expression of Tyrosinase family members, Bcl2, and other genes. The model predicts that in both Tfap2a and Kit null mutants there will be a phenotype of reduced melanocytes and that, because Tfap2a acts upstream of Kit, this phenotype will be more severe, or at least as severe as, in Tfap2a null mutants in comparison to Kit null mutants. Unexpectedly, this is not the case in zebrafish or mouse. Because many Tfap2 family members have identical DNA-binding specificity, we reasoned that another Tfap2 family member may work redundantly with Tfap2a in promoting Kit expression. We report that tfap2e is expressed in melanoblasts and melanophores in zebrafish embryos and that its orthologue, TFAP2E, is expressed in human melanocytes. We provide evidence that Tfap2e functions redundantly with Tfap2a to maintain kita expression in zebrafish embryonic melanophores. Further, we show that, in contrast to in kita mutants where embryonic melanophores appear to differentiate normally, in tfap2a/e doubly-deficient embryonic melanophores are small and under-melanized, although they retain expression of mitfa. Interestingly, forcing expression of mitfa in tfap2a/e doubly-deficient embryos partially restores melanophore differentiation. These findings reveal that Tfap2 activity, mediated redundantly by Tfap2a and Tfap2e, promotes melanophore differentiation in parallel with Mitf by an effector other than Kit. This work illustrates how analysis of single-gene mutants may fail to identify steps in a GRN that are affected by the redundant activity of related proteins.

Spontaneous melanotic lesions in axillary seabream, Pagellus acarne (Risso).

In this article, we describe spontaneous melanotic lesions in the skin of axillary seabream, Pagellus acarne (Risso), from a defined area of the Portuguese Coast, located in Cabo da Roca and Foz do Arelho. The lesions corresponded to the black pigmentation spots on the skin of the head, fins, lips and conjunctiva and, additionally, black nodules on the skin of the head and lips. In some specimens, the nodular formations in the head changed their anatomical conformation. Histologically, there were melanophores scattered along the basement membrane or forming aggregates in the dermis, infiltrating the subcutaneous tissue but not invading the adjacent muscle tissue. The aim of this study was to characterize the macroscopic and microscopic features of the pigmented lesions. These fish show sessile hyperpigmented lesions (spots) that correspond to proliferative lesions of melanophores in the dermis and nodular lesions that correspond to neoplastic lesions, melanophoromas. The melanophores in such lesions showed high concentration of melanin in the cytoplasm, moderate pleomorphism and compact distribution throughout all of the dermis.

Interactions between zebrafish pigment cells responsible for the generation of Turing patterns.

The reaction-diffusion system is one of the most studied nonlinear mechanisms that generate spatially periodic structures autonomous. On the basis of many mathematical studies using computer simulations, it is assumed that animal skin patterns are the most typical examples of the Turing pattern (stationary periodic pattern produced by the reaction-diffusion system). However, the mechanism underlying pattern formation remains unknown because the molecular or cellular basis of the phenomenon has yet to be identified. In this study, we identified the interaction network between the pigment cells of zebrafish, and showed that this interaction network possesses the properties necessary to form the Turing pattern. When the pigment cells in a restricted region were killed with laser treatment, new pigment cells developed to regenerate the striped pattern. We also found that the development and survival of the cells were influenced by the positioning of the surrounding cells. When melanophores and xanthophores were located at adjacent positions, these cells excluded one another. However, melanophores required a mass of xanthophores distributed in a more distant region for both differentiation and survival. Interestingly, the local effect of these cells is opposite to that of their effects long range. This relationship satisfies the necessary conditions required for stable pattern formation in the reaction-diffusion model. Simulation calculations for the deduced network generated wild-type pigment patterns as well as other mutant patterns. Our findings here allow further investigation of Turing pattern formation within the context of cell biology.

[Specific features of the melanophore system in different color morphs of larvae of the common toad (Bufo bufo L.)].

In a natural pond among usual black larvae of the common toad (Bufo bufo L.), a few unusual individuals of red-olive coloring were found out. In both morphs we investigated the melanophores of skin using different methods. The ESR-spectrometric analysis has shown the absence of distinctions between morphs by the amount of melanin. Analysis of total preparations of skin has shown the presence of various kinds of melanophore cells both in the derma and in the epidermis. Among typical melanophores, essentially differing cells appeared (atypical cells). In black morph tadpoles, the number of all kinds of melanophores is significantly greater than in red-olive morphs. It is shown that dark coloring is connected with a considerable number of atypical cells in the epidermis imposed on a dense layer of typical dermal melanophores with dispersed melanin.

Actin dynamics is essential for myosin-based transport of membrane organelles.

Actin filaments that serve as "rails" for the myosin-based transport of membrane organelles [1-4] continuously turn over by concurrent growth and shortening at the opposite ends [5]. Although it is known that dynamics of actin filaments is essential for many of the actin cytoskeleton functions, the role of such dynamics in myosin-mediated organelle transport was never studied before. Here, we addressed the role of turnover of actin filaments in the myosin-based transport of membrane organelles by treating cells with the drugs that suppress actin-filament dynamics and found that such a suppression significantly inhibited organelle transport along the actin filaments without inhibiting their intracellular distribution or the activity of the myosin motors. We conclude that dynamics of actin filaments is essential for myosin-based transport of membrane organelles and suggest a previously unknown role of actin-filament dynamics in providing the "rails" for continuous organelle movement resulting in the increased distances traveled by membrane organelles along the actin filaments.