Journal: eLife

Article Title: The GNU subunit of PNG kinase, the developmental regulator of mRNA translation, binds BIC-C to localize to RNP granules

doi: 10.7554/eLife.67294

Figure Lengend Snippet: ( A ) Immunoblot analysis of in vitro pull-down of GNU by BIC-C. Recombinantly expressed and purified MBP-tagged full-length GNU or GNU ΔSAM was incubated with GST-BIC-C followed by a GST pull-down. As a control, GST pull-down after incubation with MBP was performed. In contrast to the robust pull-down of MBP-GNU, only slight amounts of MBP-GNU ΔSAM were pulled down by GST-BIC-C. The levels of GST-BIC-C pulled down are comparable between all samples. ( B, C ) Fertilized embryos were collected for 2 hr from wild-type ( Oregon R ), gnu wt -gfp ( gnu wt -gfp; gnu 305 / gnu 305 ), gnu ΔSAM -gfp ( gnu ΔSAM -gfp; gnu 305 /gnu 305 ), or gnu 305 ( gnu 305 /gnu 305 ) females. Embryos were fixed and stained with DAPI. ( B ) Representative images of embryonic phenotypes. The embryos from wild-type and gnu wt -gfp mothers show normal early nuclear division cycles, whereas the gnu 305 embryo shows only a few giant nuclei. These nuclei are the consequence of DNA replication in the absence of nuclear division; the number of separate nuclei depends on whether polyploid polar bodies fuse and whether any mitotic divisions occur ( ; ). The gnu ΔSAM -gfp embryos show (from left to right panels) normal early cycles, defective blastoderm, and a few giant nuclei. Scale bar represents 100 μm. ( C ) Quantification of the embryonic phenotypes. Two independent transgenic lines were analyzed for gnu wt -gfp ( gnu wt -gfp 1-5 and gnu wt -gfp 1-8 ) and gnu ΔSAM -gfp ( gnu ΔSAM -gfp 3-2 and gnu ΔSAM -gfp 3-8 ). At least 300 embryos were scored for each transgenic line and at least 200 for the Oregon R and gnu 305 controls. ( D ) Immunoblot analysis of protein levels in mature oocytes and embryos from gnu mutants. Extracts were made from mature oocytes and embryos collected for 1 hr from Oregon R , gnu wt -gfp , gnu ΔSAM -gfp, and gnu 305 females, and the levels of GNU, CYCB, BIC-C, and ME31B were examined by immunoblot. αTUB was used as a loading control. Two independent transgenic gnu wt -gfp and gnu ΔSAM -gfp lines were examined. 30 oocytes or embryos were collected for each sample, and the equivalent of 10 oocytes was loaded into the gel per sample. Shown is one of two biological replicates. ( E ) In vitro assay of PNG kinase activity. Purified MBP-tagged GNU WT , GNU ΔSAM , or GNU P17L was incubated with the recombinant PNG kinase complex and Myelin Basic Protein (an in vitro phosphorylation target of PNG). Levels of phosphorylation of Myelin Basic Protein by PNG with radiolabeled phosphate were measured by autoradiography. MBP-GNU P17L was used as a negative control, as this amino acid change affects the ability of GNU to activate PNG kinase. In contrast, both GNU WT and GNU ΔSAM activate PNG kinase. The levels of Myelin Basic Protein are comparable across samples, as assessed by Coomassie staining. Figure 2—source data 1. Raw immunoblots from and figure with labeled bands. Figure 2—source data 2. Total embryo counts for gnu rescue experiment results in . Figure 2—source data 3. Raw immunoblots from and figure with labeled bands. Figure 2—source data 4. Raw Coomassie-stained gel and autoradiograph from . Figure with labeled bands.

Article Snippet: Recombinant DNA reagent , pCasPeR4_gnu-wt-gfp , , RRID: Addgene_113005 , .

Techniques: Western Blot, In Vitro, Purification, Incubation, Control, Staining, Transgenic Assay, Activity Assay, Recombinant, Phospho-proteomics, Autoradiography, Negative Control, Labeling

Journal: eLife

Article Title: The GNU subunit of PNG kinase, the developmental regulator of mRNA translation, binds BIC-C to localize to RNP granules

doi: 10.7554/eLife.67294

Figure Lengend Snippet:

Article Snippet: Recombinant DNA reagent , pCasPeR4_gnu-wt-gfp , , RRID: Addgene_113005 , .

Techniques: Marker, Generated, Expressing, Sterility, Labeling, Recombinant, Fluorescence, Magnetic Beads, Immunoprecipitation, Western Blot, Enzyme-linked Immunosorbent Assay, Software, Imaging

Journal: Applied Soft Computing

Article Title: A computational tool for trend analysis and forecast of the COVID-19 pandemic

doi: 10.1016/j.asoc.2021.107289

Figure Lengend Snippet: Main screen of the Graphical User Interface (GUI), with worldwide data from the European Centre for Disease Prevention and Control (ECDC). Note the list of countries on the left side.

Article Snippet: Main screen of the Graphical User Interface (GUI), with U.S. data from Johns Hopkins University.

Techniques: Control

Journal: Applied Soft Computing

Article Title: A computational tool for trend analysis and forecast of the COVID-19 pandemic

doi: 10.1016/j.asoc.2021.107289

Figure Lengend Snippet: Main screen of the Graphical User Interface (GUI), with U.S. data from Johns Hopkins University. Note the list of counties on the left side and the search for “Illinois” on the upper left corner.

Article Snippet: Main screen of the Graphical User Interface (GUI), with U.S. data from Johns Hopkins University.

Techniques: