The goals of the study were: (1) to explore the communication between human mesenchymal stem cells (MSC) and rat cardiac myocytes resulting in differentiation of the stem cells and, (2) to evaluate the role of mitochondria in it. Light and fluorescence microscopy as well as scanning electron microscopy revealed that after co-cultivation, cells formed intercellular contacts and transient exchange with cytosolic elements could be observed. The transport of cytosolic entity had no specific direction. Noticeably, mitochondria also could be transferred to the recipient cells in a unidirectional fashion (towards cardiomyocytes only). Transmission electron microscopy revealed significant variability in both the diameter of intercellular contacting tubes and their shape. Inside of these nanotubes mitochondria-resembling structures were identified. Moreover, after co-cultivation with cardiomyocytes, expression of human-specific myosin was revealed in MSC. Thus, we speculate that: (1) transport of intracellular elements to MSC possibly can determine the direction of their differentiation and, (2) mitochondria may be involved in the mechanism of the stem cell differentiation. It looks plausible that mitochondrial transfer to recipient cardiomyocytes may be involved in the mechanism of failed myocardium repair after stem cells transplantation.

Some picornaviruses, for example, poliovirus, increase bidirectional permeability of the nuclear envelope and suppress active nucleocytoplasmic transport. These activities require the viral protease 2Apro . Here, we studied nucleocytoplasmic traffic in cells infected with encephalomyocarditis virus (EMCV; a cardiovirus), which lacks the poliovirus 2A pro -related protein. EMCV similarly enhanced bidirectional nucleocytoplasmic traffic. By using the fluorescent "Timer" protein, which contains a nuclear localization signal, we showed that the cytoplasmic accumulation of nuclear proteins in infected cells was largely due to the nuclear efflux of "old" proteins rather than impaired active nuclear import of newly synthesized molecules. The nuclear envelope of digitonin-treated EMCV-infected cells permitted rapid efflux of a nuclear marker protein. Inhibitors of poliovirus 2A pro did not prevent the EMCV-induced efflux. Extracts from EMCV-infected cells and products of in vitro translation of viral RNAs contained an activity increasing permeability of the nuclear envelope of uninfected cells. This activity depended on the expression of the viral leader protein. Mutations disrupting the zinc finger motif of this protein abolished its efflux-inducing ability. Inactivation of the L protein phosphorylation site (Thr473Ala) resulted in a delayed efflux, while a phosphorylation-mimicking (Thr473Asp) replacement did not significantly impair the efflux-inducing ability. Such activity of extracts from EMCV-infected cells was suppressed by the protein kinase inhibitor staurosporine. As evidenced by electron microscopy, cardiovirus infection resulted in alteration of the nuclear pores, but it did not trigger degradation of the nucleoporins known to be degraded in the poliovirus-infected cells. Thus, two groups of picornaviruses, enteroviruses and cardioviruses, similarly alter the nucleocytoplasmic traffic but achieve this by strikingly different mechanisms.Picornaviruses, small nonenveloped icosahedral animal viruses with a single-stranded RNA genome of positive (mRNA) polarity, encompass the Enterovirus, Rhinoviruses, Cardiovirus, Aphthovirus, Parechovirus, and some other genera (64). All essential steps of their reproduction, such as translation, RNA synthesis, and encapsidation, take place in the cytoplasm of infected cells. The nonessential role of the nucleus for their reproduction follows from their ability to fulfill the complete infectious cycle in nuclei-free cytoplasts (31, 60) or cytoplasmic extracts (7,52,71). This fact, however, does not mean that the nuclei are not involved in the infectious process. Indeed, virusspecific proteins have been detected in the nuclei of poliovirusinfected (11, 29) and encephalomyocarditis virus (EMCV)-infected (5, 6) cells. Poliovirus proteases 2A and 3C are known to target a variety of nuclear transcription factors and histones (66,78,79,80). The EMCV 2A protein enters the nucleoli and interacts there with a ribosome precursor, contributing thereby to alterations in the translation c...

The nucleus contains distinct nuclear bodies (NBs); nucleolus is the largest and the most studied NB, but its role in the functioning of the nucleus is far from being fully understood. The nucleolus is not surrounded by a membrane, yet it contains DNA, RNA and a set of proteins that can either be retained in the nucleolus or rapidly shuttle between the nucleoplasm, the nucleolus and the cytoplasm in response to various stimuli. The emerging evidence points to the central function of the nucleolus in organizing many nuclear functions besides RNA polymerase I transcription and ribosome biogenesis. Here we discuss the functions of the nucleolus related to the shuttling of proteins and nucleic acids between nucleolus and nucleoplasm. The functional processes affected by shuttling of nucleolar components include 3D organization of the genome, stress response, DNA repair and recombination, transcription regulation, telomere maintenance and other essential cellular functions. Powered by Editorial Manager® and ProduXion Manager® from Aries Systems Corporation Highlights  Nucleolus is a PolI transcription factory and a place of ribosome assembly, but is also performs many other functions that are commonly referred to as non-canonical functions of nucleolus  Nucleolus organizes the adjacent chromatin into a large-scale repressive hub underlying the spatial segregation of active and repressive chromatin compartments.  The interphase chromosomes are attached to the nucleolus via nucleolus-associated domains (NADs).  Protein shuttling between the nucleolus and the nucleoplasm regulates a multitude of nuclear processes including DNA repair, recombination, transcription and telomere maintenance.

Representatives of several picornavirus genera have been shown previously to significantly enhance noncontrollable bidirectional exchange of proteins between nuclei and cytoplasm. In enteroviruses and rhinoviruses, enhanced permeabilization of the nuclear pores appears to be primarily due to proteolytic degradation of some nucleoporins (protein components of the pore), whereas this effect in cardiovirus-infected cells is triggered by the leader (L) protein, devoid of any enzymatic activities. Here, we present evidence that expression of L alone was sufficient to cause permeabilization of the nuclear envelope in HeLa cells. In contrast to poliovirus, mengovirus infection of these cells did not elicit loss of nucleoporins Nup62 and Nup153 from the nuclear pore complex. Instead, nuclear envelope permeabilization was accompanied by hyperphosphorylation of Nup62 in cells infected with wild-type mengovirus, whereas both of these alterations were suppressed in L-deficient virus mutants. Since phosphorylation of Nup62 (although less prominent) did accompany permeabilization of the nuclear envelope prior to its mitotic disassembly in uninfected cells, we hypothesize that cardiovirus L alters the nucleocytoplasmic traffic by hijacking some components of the normal cell division machinery. The variability and biological significance of picornaviral interactions with the nucleocytoplasmic transport in the infected cells are discussed.Productive viral interaction with host cells requires generation of an intracellular environment ensuring efficient synthesis and assembly of virally encoded macromolecules. A fundamental aspect of this process is overcoming a variety of host defensive tools, collectively named innate immunity. Some of the viral antidefensive measures are direct results of enzymatic activities of viral proteins, as exemplified by proteolytic degradation of host translation or transcription factors. However, the viral antidefensive armament in the case of small-genome viruses is usually relatively limited, and therefore such viruses have evolved the capacity to divert ("hijack") cellular proteins, pathways, and structures to perform proviral functions. A group of viruses relatively well studied in this respect, is represented by picornaviruses-small, nonenveloped animal viruses with a 7.2-to 8.5-kb RNA genome of positive polarity (46).The Picornaviridae family is composed of at least nine genera (48), which share general genome organization but differ from each other in some important aspects such as, for example, the capsid structure (and consequently the usage of cellular receptors), the structure of replicative and translational cis elements, and the presence or absence of structurally and functionally unrelated accessory (usually nonessential) proteins (2).In recent years, we have studied, among other activities exhibited by picornaviruses, their ability to trigger bidirectional permeabilization of the nuclear envelope, facilitating noncontrollable exchange of macromolecules between the nucleus and cytoplasm...

Apoptosis is a common antiviral defensive mechanism that potentially limits viral reproduction and spread. Many viruses possess apoptosis-suppressing tools. Here, we show that the productive infection of HeLa cells with encephalomyocarditis virus (a cardiovirus) was not accompanied by full-fledged apoptosis (although the activation of caspases was detected late in infection) but rather elicited a strong antiapoptotic state, as evidenced by the resistance of infected cells to viral and nonviral apoptosis inducers. The development of the antiapoptotic state appeared to depend on a function(s) of the viral leader (L) protein, since its mutational inactivation resulted in the efflux of cytochrome c from mitochondria, the early activation of caspases, and the appearance of morphological and biochemical signs of apoptosis in a significant proportion of infected cells. Infection with both wild-type and L-deficient viruses induced the fragmentation of mitochondria, which in the former case was not accompanied with cytochrome c efflux. Although the exact nature of the antiapoptotic function(s) of cardioviruses remains obscure, our results suggested that it includes previously undescribed mechanisms operating upstream and possibly downstream of the mitochondrial level, and that L is involved in the control of these mechanisms. We propose that cardiovirus L belongs to a class of viral proteins, dubbed here security proteins, whose roles consist solely, or largely, in counteracting host antidefenses. Unrelated L proteins of other picornaviruses as well as their highly variable 2A proteins also may be security proteins. These proteins appear to be independent acquisitions in the evolution of picornaviruses, implying multiple cases of functional (though not structural) convergence.