Journal: Nature

Article Title: Cytoplasmic control of intranuclear polarity by human cytomegalovirus

doi: 10.1038/s41586-020-2714-x

Figure Lengend Snippet: a, Emerin is polarized in HCMV-infected cells. Lines represent mean ± SEM; n= 19,428 cells total from 3 independent biological replicates. b, Onset of Emerin polarization and formation of nuclear F-actin is detectable by 24h, as nuclear rotation begins. Emerin polarity is established over the nuclear rotation period and is sustained, while F-actin dissipates after the rotation phase. c-d, Stills from - showing nuclear F-actin formation during mock or HCMV (TB40/E-UL99-mCherry) infection of NHDFs expressing nuclear actin chromobody (nAC-TagGFP). Early and late stages of infection are shown in c and d, respectively. e , Expression of actin-binding mutants (m151, m175) of Emerin impairs nuclear F-actin formation. Representative images and quantification of nuclear F-actin (nAC) frequency are shown for each condition; bars represent mean ± SEM, n = 1,230 cells total, **p≤0.01, two-tailed student’s t-test. All data shown is representative of 3 independent biological replicates.

Article Snippet: The viral vector encoding a nuclear actin chromobody (actin targeting nanobody tagged with TagGFP2 and a nuclear localization signal; Chromotek: acg-n) was generated by cloning the chromobody sequence into the pWPXL lentiviral vector (Addgene: #12257) using nAC-for (5’-CGCACCGGTGGATCCGCCACCATGGCTCAGGTGCAGCTGGTGG-3’) and nAC-rev (5’-CCGGAATTCACTAGTTTATTACACCTTCCGCTTTTTCTT-3’) primers to generate pWPXL-TagGFP2-nAC, as previously described .

Techniques: Infection, Expressing, Binding Assay, Two Tailed Test