Characterization of a bicistronic knock-in reporter mouse model for investigating the role of CABLES2 in vivo

Two members of the CDK5 and ABL enzyme substrate (CABLES) family, CABLES1 and CABLES2, share a highly homologous C-terminus. They interact and associate with cyclin-dependent kinase 3 (CDK3), CDK5, and c-ABL. CABLES1 mediates tumor suppression, regulates cell proliferation, and prevents protein degradation. Although Cables2 is ubiquitously expressed in adult mouse tissues at RNA level, the role of CABLES2 in vivo remains unknown. Here, we generated bicistronic Cables2 knock-in reporter mice that expressed CABLES2 tagged with 3×FLAG and 2A-mediated fluorescent reporter tdTomato. Cables2-3×FLAG-2A-tdTomato (Cables2^Tom) mice confirmed the expression of Cables2 in various mouse tissues. Interestingly, high intensity of tdTomato fluorescence was observed in the brain, testis and ovary, especially in the corpus luteum. Furthermore, immunoprecipitation analysis using the brain and testis in Cables2^Tom/Tom revealed interaction of CABLES2 with CDK5. Collectively, our new Cables2 knock-in reporter model will enable the comprehensive analysis of in vivo CABLES2 function.     

Antiviral activities of mycophenolic acid and IMD-0354 against SARS-CoV-2

In this study, the anti–severe acute respiratory syndrome coronavirus-2 (anti-SARS-CoV-2) activity of mycophenolic acid (MPA) and IMD-0354 was analyzed. These compounds were chosen based on their antiviral activities against other coronaviruses. Because they also inhibit dengue virus (DENV) infection, other anti-DENV compounds/drugs were also assessed. On SARS-CoV-2-infected VeroE6/TMPRSS2 monolayers, both MPA and IMD-0354, but not other anti-DENV compounds/drugs, showed significant anti-SARS-CoV-2 activity. Although MPA reduced the viral RNA level by only approximately 100-fold, its half maximal effective concentration was as low as 0.87 µ m , which is easily achievable at therapeutic doses of mycophenolate mofetil. MPA targets the coronaviral papain-like protease and an in-depth study on its mechanism of action would be useful in the development of novel anti-SARS-CoV-2 drugs.     

Determination of two different types of cellular cementogenesis in rat molars: a histological and immunohistochemical study.

To elucidate the attachment mechanism of dentin and cellular cementum, developing and developed cellular cementum of rat molars was examined by light microscopy. Routine histological staining, immunohistochemical staining for bone sialoprotein (BSP) and osteopontin (OPN), and digestion tests with trypsin were conducted. Two different types of cellular cementogenesis were established, one on the mesial (type I cementogenesis) and one on the distal sides (type II cementogenesis) of the examined roots. In the type I cementogenesis a thin initial cementum layer, which was fibril-poor, hematoxylin-stained, and immunopositive for BSP and OPN, appeared on the mineralized dentin. With cellular cementogenesis, the layer became the cemento-dentinal junction. The cementum mineralization did not precede the dentin mineralization. After trypsin treatment the cemento-dentinal junction lost immunoreactivity for BSP and OPN and the cementum was detached from the dentin. In the type II cementogenesis the cellular cementum formed directly on the predentin without the initial cementum layer and the cementum mineralization preceded the dentin mineralization. Cemental and predentinal fibrils appeared to intermingle, as the cemento-dentinal junction was indiscernible by any staining. Trypsin treatment did not cause cementum detachment. The findings of the present study suggest that: (1) The type I cementogenesis requires the intervening initial cementum to bind cementum and dentin and to induce the cementum mineralization. (2) In the type II cementogenesis the cemento-dentinal attachment depends on fibril intermingling and the cementum mineralization advances apically and very rapidly, probably producing mineralization foci. (3) The formation of the initial cementum depends on the speed of the cementogenesis in the apical direction.     

Involvement of the 3’ Untranslated Region in Encapsidation of the Hepatitis C Virus

Although information regarding morphogenesis of the hepatitis C virus (HCV) is accumulating, the mechanism(s) by which the HCV genome encapsidated remains unknown. In the present study, in cell cultures producing HCV, the molecular ratios of 3’ end- to 5’ end-regions of the viral RNA population in the culture medium were markedly higher than those in the cells, and the ratio was highest in the virion-rich fraction. The interaction of the 3’ untranslated region (UTR) with Core in vitro was stronger than that of the interaction of other stable RNA structure elements across the HCV genome. A foreign gene flanked by the 3’ UTR was encapsidated by supplying both viral NS3-NS5B proteins and Core-NS2 in trans. Mutations within the conserved stem-loops of the 3’ UTR were observed to dramatically diminish packaging efficiency, suggesting that the conserved apical motifs of the 3´ X region are important for HCV genome packaging. This study provides evidence of selective packaging of the HCV genome into viral particles and identified that the 3’ UTR acts as a cis-acting element for encapsidation.     

Demonstration of bacterium-free very rapid reverse genetics system using mumps virus

The reverse genetics system is a very powerful tool for analyzing the molecular mechanisms of viral propagation and pathogenesis. However, full-length genome plasmid construction is highly time-consuming and laborious, and undesired mutations may be introduced by Escherichia coli. This study shows a very rapid E. coli-free method of full-genome construction using the mumps virus as an example. This method was able to reduce dramatically the time for full-genome construction, which was used very efficiently for virus rescue, from several days or more to ~2 days, with a similar accuracy and yield to the conventional method using E. coli/plasmid.     

Chemical Structure and Acaricidal Activity of 3-Alkylthiophenyl 4-Nitrophenyl Ethers

Many 4-nitro diphenylether derivatives with R–S (O)_x-substituents were systematically synthesized and their pesticidal activities were investigated. 3-n-Propylthiophenyl 4-nitro-phenyl ether and its sulfoxide analog exhibited remarkable acaricidal and chlorosis-inducing activities on the young leaves of plants. For all compounds with the substituent at the 4 position in the 3-n-propylthio (and -sulfinyl) phenyl group, chlorosis-inducing activity vanished and an acaricidal activity stronger than that of the parent compound appeared. Evidence that the acaricidal and chlorosis-inducing activities are greatly influenced by the size of the 4-sub-stituent and the hydrophobicity of R in the 3-R-S (O)_x-substituent is presented.