Journal: Nature microbiology

Article Title: TRIM23 mediates virus-induced autophagy via activation of TBK1

doi: 10.1038/s41564-017-0017-2

Figure Lengend Snippet: a, Summary of the TRIM cDNA screen (shown in ) in which 61 TRIM proteins were tested for their ability to induce GFP-LC3B puncta formation in transiently transfected HeLa cells. TRIM proteins were categorized into ‘high inducers’ of autophagy (red; defined as inducing more GFP-LC3B puncta than rapamycin stimulation), or ‘low/intermediate inducers’ and ‘non-inducers’ (magenta/blue; defined as inducing fewer GFP-LC3B puncta than rapamycin but more than empty vector transfection, or similar to vector transfection, respectively). b, Representative laser-scanning confocal microscopy images of GFP-LC3B puncta formation in HeLa cells for the top 7 hits from the TRIM cDNA screen shown in (a). Representative images for GFP-LC3B puncta formation induced by rapamycin treatment or empty vector transfection (controls) are also shown. Scale bar, 20 µm. c–e, GFP-LC3B puncta formation in HeLa cells transiently transfected with non-targeting control siRNA (NT) or siRNAs targeting the indicated TRIM proteins and subsequently infected with mutHSV-1 (MOI 4) for 8 h (c), IAV (MOI 5) for 14 h (d), or EMCV (MOI 100) for 6 h (e). Box and whisker plots show the distribution of the area of GFP-LC3B for 3 individual images, each containing ~30 cells. f, Summary of the results of GFP-LC3B puncta formation from the RNAi screen using viral stimuli (c-e) or rapamycin treatment . NV, no infection. *p<0.05, ***p<0.0005 (ANOVA test). Data are representative of one screen ( a ), or two ( b ) or three ( c–f ) independent experiments.

Article Snippet: RSV (A2 strain) and EMCV (EMC strain) were purchased from ATCC.

Techniques: Transfection, Plasmid Preparation, Confocal Microscopy, Control, Infection, Whisker Assay

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: A . Typical structure for the genomic RNA of EMCV. The magenta double head arrow indicates the PCR target region to detect a reverse transcribed EMCV genome. B . The illustration shows a strategy of the testis culture and viral infection. A reverse-transcription of the viral genomic RNA is evaluated by PCR and electrophoresis. C . A model for the pathway from viral infection to endogenization. A genomic sequence of an RNA virus is considered to be reverse-transcribed by host-derived reverse-transcriptase. Subsequently, the cDNA might be integrated into the host genome by non-homologous end joining. D . The photographs show typical images for transverse section of seminiferous tubule in the cultured testes at 21 dpi and the uninfected control. Arrowheads indicate spermatogonia. Scale bars, 20 µm. E . The electrophoresis shows a result of PCR to detect DNA fragment that is identical to the EMCV genome. The samples were harvested from the EMCV-infected testes at 1-, 3-, 5-, 7-, 14-, and 21-dpi. Expected size of the amplicon is 315 bp. M, size marker. EI, EMCV-infected. UI, uninfected. PC, positive control (EMCV cDNA). NC, negative control (empty vector).

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Reverse Transcription, Infection, Electrophoresis, Sequencing, Virus, Derivative Assay, Non-Homologous End Joining, Cell Culture, Control, Amplification, Marker, Positive Control, Negative Control, Plasmid Preparation

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: The electrophoresis shows a result of PCR to detect DNA fragment that is identical to the EMCV genome. The samples were harvested from the EMCV-infected TERT (+/+) or TERT (−/−) littermate testes at 5- and 7-dpi. Expected size of the amplicon is 315 bp. M, size marker. NC, negative control (uninfected testis).

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Electrophoresis, Infection, Amplification, Marker, Negative Control

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: A . The illustration shows a strategy of the viral infection to live animal and a molecular hypothesis for subsequent reverse-transcription and endogenization. B . The line graph indicates a survival rate of the EMCV-infected adult male mice. All infected mice were dead within 4 dpi except lowest viral titer (1.0 × 10 −1 pfu). C . The electrophoresis shows a result of PCR to detect DNA fragment that is identical to the EMCV genome. The DNA samples were harvested from testes extracted from the male mice deceased after EMCV injection at the titer of 10 × 10 1 , 10 3 , and 10 5 pfu. Expected size of the amplicon is 315 bp. M, size marker. D . The electrophoresis shows a result of PCR to detect DNA fragment that is identical to the EMCV genome. The DNA samples were harvested from mating plugs derived from the male mice survived after EMCV injection at the titer of 10 × 10 −1 pfu. Expected size of the amplicon is 315 bp. M, size marker. NC, negative control (empty vector). PC, positive control (EMCV cDNA).

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Infection, Reverse Transcription, Electrophoresis, Injection, Amplification, Marker, Derivative Assay, Negative Control, Plasmid Preparation, Positive Control

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: The illustration shows the time course of the EMCV infection to mice and the actual timing for the important events and sampling in this study. The events described above are about the infection and mating. The others are samplings mentioned in the main text and figures.

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Infection, Sampling

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: A . The illustration shows a strategy to validate a heredity of the endogenized EMCV sequence from the infected male to the offspring. B . The line graph indicates a survival rate of F1 offspring obtained from the mating between the survived EMCV-infected male and uninfected female. Each color means the littermate derived from respective male mice. C . The electrophoresis shows a result of PCR to detect an endogenized EMCV fragment in the lethal offspring. The DNA samples were harvested from livers extracted from the F1 neonate deceased within 3 d after birth. The numbers were assigned for the parental male (#1∼#3) and littermate recognition. Expected size of the amplicon is 315 bp. M, size marker. NC, negative control (empty vector). PC, positive control (EMCV cDNA). D . The electrophoresis shows a result of PCR to detect an endogenized EMCV fragment in the survived offspring. The DNA samples were harvested from earlobes cut from the F1 offspring and their parent males. The numbers were assigned for the parental male (#1∼#4) and littermate recognition. Expected size of the amplicon is 315 bp. NC, negative control (empty vector). PC, positive control (EMCV cDNA). M, size marker.

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Sequencing, Infection, Derivative Assay, Electrophoresis, Amplification, Marker, Negative Control, Plasmid Preparation, Positive Control

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: The table shows the numerical data for survival rate of the F1 generation mice obtained from the mating between the EMCV-infected male and uninfected female. This figure is related to . Survival number was recorded every day until 15 days after the birth.

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Infection

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: The alignments illustration shows the raw sequence reads mapped to the EMCV and mouse genome. The genomic DNA extracted from the EMCV-infected testes was used for the NGS. The above read contains a small insertion in the border of the EMCV and mouse sequences. The other is considered to have undergone a precise non-homologous end joining, while an extra sequence was observed in the terminals of the read.

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Sequencing, Infection, Non-Homologous End Joining

Journal: bioRxiv

Article Title: Heritable endogenization of an RNA virus in a mammalian species

doi: 10.1101/2020.01.19.911933

Figure Lengend Snippet: A-C. The table and graphs show the body weight of the survived F1 generation mice measured at 6-weeks. The offspring of the EMCV-infected male mice tended to exhibit smaller weight than the average of the C57BL/6J strain (Male, 21.9 ± 1.8. Female, 18.5 ± 0.9.). The dotted lines in B and C indicate the averages in each sex. The data were obtained from the Jackson Laboratory website ( https://www.jax.org/jax-mice-and-services/strain-data-sheet-pages/body-weight-chart-000664 ).

Article Snippet: The EMCV strain NIID-NU1 (DDBJ, #LC508268) was propagated in Vero cells (JCRB9013) purchased from the Japanese Collection of Research Bioresources (JCRB; Ibaraki, Japan).

Techniques: Infection