[en] Highlights: • Vimentin intermediate filaments interact with caveolin-1. • In cytoplasm, where there is pronounced vimentin network, there is less caveolin-1 positive vesicles. • Caveolin-1 positive vesicles show less mobility in vimentin-enriched area. • Vimentin depletion do not affect the protein expression levels of caveolae components. • Vimentin depletion affects the mobility distribution of the cytoplasmic caveolin-1. Both the cytoskeletal intermediate filaments (IFs) and cytoplasmic caveolae contribute to active processes such as cell migration, morphogenesis and vesicular trafficking, but the interplay between these two systems has remained elusive. Here, we find that vimentin and nestin IFs interact with caveolae central component caveolin-1 (CAV-1) and importantly, restrain the intracellular trafficking of CAV-1 positive vesicles by serving as a physical barrier. Consequently, CAV-1 vesicles show less density and mobility in vimentin IFs enriched region, which is a substrate stiffness independent process. Moreover, depletion of vimentin IFs releases the slow movement proportion of CAV-1 positive vesicles and thus increases their cytoplasmic dynamics, whereas the expression of caveolae-associated protein CAV-1, CAV-2 and cavin-1 were unaffected. Collectively, these results reveal a negative role of IFs in regulating the trafficking of intracellular CAV-1 vesicles in live cells.

[en] Highlights: • Indirubin 3′-oxime inhibited anticancer agent-induced nuclear import of YB-1 in HepG2 cells. • Actinomycin D-activated MDR1 expression was suppressed in the presence of indirubin 3′-oxime. • An increase of drug sensitivity was observed in HepG2 cells pretreated with indirubin 3′-oxime. Hepatocellular carcinoma (HCC) is a disease with poor prognosis. Nuclear accumulation of YB-1 is closely related to the malignancy of HCC. Treatment with anticancer agents often induces translocation of YB-1 from cytoplasm to nucleus and activates the expression of multidrug resistance gene 1 (MDR1). Therefore, any effective inhibitor of this phenomenon would be useful for cancer treatment. Here we examined various indirubin derivatives and found that indirubin 3′-oxime inhibits actinomycin D-induced nuclear transport of YB-1 and suppresses the activation of MDR1 gene expression in the human hepatocellular carcinoma cell line HepG2. Furthermore, use of both indirubin 3′-oxime and actinomycin D in combination increased the anticancer effect on HepG2 cells. Indirubin 3′-oxime is a novel and efficient inhibitor of anticancer agent-induced YB-1 nuclear translocation.

[en] Highlights: • Defects in desmosome formation observed upon 14-3-3γ loss are corrected by targeting PG to the cell border. • The restoration of desmosome formation by Plakoglobin in the absence of 14-3-3γ requires PKP3. • 14-3-3γ regulates desmosome formation via multiple pathways in different cell types. Desmosomes are cell-cell adhesion junctions that anchor intermediate filaments. Loss of 14-3-3γ in HCT116 cells led to defects in desmosome assembly due to a decrease in the transport of Plakoglobin (PG) to the cell border thus disrupting desmosome formation. Desmosome formation in cells lacking 14-3-3γ was restored by artificially localizing PG to the cell border by fusing it to EGFP-f (PG-EGFP-f). These results suggest that a major role of 14-3-3γ in desmosome assembly is to transport PG to the cell border leading to the initiation of desmosome formation.

[en] Proteasomes are the major actors of nonlysosomal cytoplasmic protein degradation. In particular, these large protein complexes (about 2500 kDa) are considered to be responsible for muscular degradation during skeletal muscle atrophy. Despite their unusual and important size, they are widely described as soluble and mobile in the cytoplasm. In mature skeletal muscle, we have previously observed a sarcomeric distribution of proteasomes, as revealed by the distribution of α1/p27K, a subunit of the 20S core-particle (prosome) of proteasome. Here, we extend these observations at the electron microscopic level in vivo. We also show that this sarcomeric pattern is dependent of the extension of the sarcomere. Using isolated myofibrils, we demonstrate that proteasomes are still attached to the myofibrils after the isolation procedure, and reproduce the observations made in vivo. In addition, the extraction of actin by gelsolin largely removes proteasomes from isolated myofibrils, but some of them are held in place after this extraction, showing a sarcomeric disposition in the absence of any detectable actin, and suggesting the existence of another molecular partner for these interactions. From these results, we conclude that most of detectable 20S proteasomes in skeletal muscle cells is tightly attached to the myofibrils

[en] The deduced amino acid (aa) sequence of L2 of the newly sequenced HPV51 strain, isolated by Matsukura and Sugase (Ma-strain), was markedly different from that of the prototype HPV51 isolated by Nuovo et al. (Nu-strain) (GenBank (M62877)) in two regions: aa 95-99 (region I) and aa 179-186 (region II). The two regions of Ma-strain were homologous to those of the other mucosal HPVs. The aa sequences of the N-terminal and C-terminal regions of Ma-L2 and Nu-L2 were identical and contained the nuclear localizing signal (NLS). When expressed in HEK293 cells, Ma-strain L2 (Ma-L2) was located in the nucleus but Nu-strain L2 (Nu-L2), in the cytoplasm. The chimeric L2s having both Nu-L2 regions I and II were located in the cytoplasm, and those having one of them were located both in the nucleus and cytoplasm, suggesting that Nu-L2 regions I and II inhibit the NLS function. For a better understanding of a role of L2 in infection, pseudovirion (PV) preparations were produced with a reporter, Ma-strain L1, and various L2s (Ma-L2, Nu-L2, or the chimeric L2s). These PV preparations contained structurally similar particles composed of L1 and L2 and the packaged reporter plasmid at a similar level. The reporter expression was not induced in HEK293 cells after inoculation with PVs containing the L2s that are incapable of localizing in the nucleus when expressed alone. Among PVs containing L2s capable of localizing in the nucleus, the reporter expression was induced only by PVs containing Ma-L2 region I. Thus, the results indicate that the expression of the reporter in the HPV51 PV requires the nuclear localizing ability of L2 and another unknown function associated with region I.

[en] In this paper, we study the impacts of an initial axial force and the surface effects on the dynamic characteristics of an embedded bioliquid filled microtubule (MT) under a walking motor protein. We model the microtubule using the orthotropic Timoshenko Beam (TB) model. The impact of the surrounding cytoplasm is considered by modelling it as a two parameters Pasternak model. We used the modified couple stress theory (MCST) in order to include small-scale effects to derive the motion equations by applying Hamilton’s principle for the orthotropic TB model. The influence of motor protein motion, surrounding cytoplasm matrix, surface effects and the initial axial loading are examined for the deflection, rotation and coupled vibration frequency of the embedded microtubule simply supported on both ends. We found that the initial axial loading and the magnitude of the moving load due to motor protein motion renders the simply supported microtubule less stable. It is seen that surface parameters tend to increase the stability of the microtubule as they enhance the frequency of vibration and reduce the normalised dynamic deflection and rotation for all vibration modes. The Winkler and Pasternak parameters are found to greatly affect the dynamic behaviour of the liquid filled microtubules. (author)

[en] Highlights: • The NLRP3 inflammasome activation is attenuated by PSMD10, BCL2L10, CTSS, and ILF2. • ILF2 represses inflammasome activation, pro-Casp-1 cleavege, and pro-IL-1β maturation. • ILF2 interacts with NLRP3 to inhibit the inflammasome activation in the cytoplasm. • ILF2 represses ATP-induced activation of endogenous NLRP3 inflammasome in macrophages. The activation of the NLRP3 inflammasome is a key process of host immune response that establishes the first line of defense against pathogen infections and cellular stresses, whereas excessive inflammasome activation may damage the hosts, and thus it must be precisely controlled. However, the mechanism underlying the repression of the NLRP3 inflammasome activation remains largely unknown. In this study, by establishing and using a reconstructed NLRP3 inflammasome activation system, we reveal that the NLRP3 inflammasome activation, pro-caspase-1 cleavage, and pro-interleukin-1β (pro-IL-1β) activation are repressed by the interleukin-enhanced binding factor 2 (ILF2). Further studies demonstrate that ILF2 represses the activation of NLRP3 inflammasome through interacting with the NACHT-associated domain (NAD) of NLRP3 and co-localized with NLRP3 in the cytoplasm of HEK293T cells. Finally, by generating a THP-1 cell line stably expressing ILF2 protein using the lentivirus infection system, we demonstrate that ILF2 represses ATP-induced activation of endogenous NLRP3 inflammasome in macrophages. Therefore, this study identifies a new role of ILF2 in the regulation of the NLRP3 inflammasome, and reveals a unique mechanism underlying the repression of the NLRP3 inflammasome activation.

[en] Highlights: • Porcine reproductive and respiratory syndrome (PRRS) affects industrial pig farming. • N protein is the main protein of PRRS virus (PRRSV) in infected cells which is important for PRRSV replication. • Moloney leukemia virus 10 protein (MOV10) is a new host restriction factor of PRRSV. • MOV10 interacts with PRRSV N protein and inhibits PRRSV replication in vitro. • MOV10 inhibits nuclear entry of N protein, thus impairing viral life cycle. Porcine reproductive and respiratory syndrome virus (PRRSV) has been a major threat to global industrial pig farming ever since its emergence in the late 1980s. Identification of sustainable and effective control measures against PRRSV transmission is a pressing problem. The nucleocapsid (N) protein of PRRSV is specifically localized in the cytoplasm and nucleus of virus-infected cells which is important for PRRSV replication. In the current study, a new host restricted factor, Moloney leukemia virus 10-like protein (MOV10), was identified as an inhibitor of PRRSV replication. N protein levels and viral replication were significantly reduced in Marc-145 cells stably overexpressing MOV10 compared with those in wild-type Marc-145 cells. Adsorption experiments revealed that MOV10 did not affect the attachment and internalization of PRRSV. Co-immunoprecipitation and immunofluorescence co-localization analyses showed that MOV10 interacted and co-localized with the PRRSV N protein in the cytoplasm. Notably, MOV10 affected the distribution of N protein in the cytoplasm and nucleus, leading to the retention of N protein in the former. Taken together, these findings demonstrate for the first time that MOV10 inhibits PRRSV replication by restricting the nuclear import of N protein. These observations have great implications for the development of anti-PRRSV drugs and provide new insight into the role of N protein in PRRSV biology.

[en] In this study, karyotype criteria of 27 accessions (17 taxa) of the two tribes Asclepiadeae (20 accessions) and Ceropegieae (7 accessions) from Egypt and Saudi Arabia belonging to subfamily Asclepiadoideae are described and polyploid variations are also discussed. Detailed karyotype features, i.e. total chromosome length (TCL), mean chromosome length (MCL) and karyotype asymmetry expressed as arm ratio (MAR), total form percent (TF %), intrachromosomal asymmetry (A1) and interchromosomal asymmetry (A2), are also described. Karyotype features of the studied accessions were used to assess the tribal relationships within the subfamily Asclepiadoideae to differentiate between taxa that belonging to the tribes Asclepiadeae and Ceropegieae in the light of the current systems of classification. In addition, new chromosome counts of 16 taxa or accessions are reported here for the first time. (author)

[en] Highlights: • A de novo mutation was identified in OLFML1 in a patient with congenital scoliosis. • Olfml1 is strongly expressed in calvarial osteoblasts in mice. • Knockdown of Olfml1 accelerates osteoblast mineralization. • Olfml1 knockdown induces nuclear translocation of YAP and activates Hippo signaling. • Olfml1 expression vector transfection retains YAP in the cytoplasm. Congenital scoliosis is a lateral curvature of the spine that is due to the presence of vertebral anomalies. Although genetic and environmental factors are involved in the pathogenesis of congenital scoliosis, the specific cause of only a small number of individuals has been identified to date. We identified a de novo missense mutation in the olfactomedin-like 1 (OLFML1) gene by whole-exome sequencing of a patient with congenital scoliosis. Then, we carried out further functional investigation in mice. An assessment of the tissue distribution of Olfml1 revealed it to be prominently expressed in developing skeletal tissues, specifically osteoblasts. Short hairpin RNA-mediated knockdown of Olfml1 in osteoblasts induced the translocation of Yes-associated protein (YAP) transcriptional coactivator from the cytoplasm to the nucleus, which accelerated the Hippo signaling pathway to promote osteoblast mineralization. In contrast, experimentally induced gain of function of Olfml1 retained YAP in the cytoplasm. There appears to exist a novel cell-autonomous mechanism by which osteoblasts avoid excess mineralization through Olfml1. Our results also indicate that mutation of OLFML1 leads to impaired osteoblast differentiation and abnormal development of bone tissue.