Our data clearly demonstrate that TRN-SR2 is the nuclear-import factor of HIV.

Human T cell lymphotropic virus I (HTLV-I) is the etiological agent for adult T cell leukemia and tropical spastic paraparesis (also termed HTLV-I-associated myelopathy). HTLV-I-infected peripheral blood T cells exhibit an initial phase of interleukin-2 (IL-2)-dependent growth; over time, by an unknown mechanism, the cells become IL-2-independent. Whereas the Jak kinases Jak1 and Jak3 and the signal transducer and activator of transcription proteins Stat3 and Stat5 are activated in normal T cells in response to IL-2, this signaling pathway was constitutively activated in HTLV-I-transformed cells. In HTLV-I-infected cord blood lymphocytes, the transition from IL-2-dependent to IL-2-independent growth correlated with the acquisition of a constitutively activated Jak-STAT pathway, which suggests that this pathway participates in HTLV-I-mediated T cell transformation.

Despite the utility of CRISPR-Cas9 nucleases for genome editing, the potential for off-target activity limits their application, especially for therapeutic purposes1,2. We developed a yeast-based assay to identify optimized Streptococcus pyogenes Cas9 (SpCas9) variants that enables simultaneous evaluation of on- and off-target activity. We screened a library of SpCas9 variants carrying random mutations in the REC3 domain and identified mutations that increased editing accuracy whilst maintaining editing efficiency. We combined four beneficial mutations to generate evoCas9, a variant that has fidelity exceeding both wild-type (79-fold improvement) and rationally designed Cas9 variants3,4 (4-fold average improvement), while maintaining near wild-type on-target editing efficiency (90% median residual activity). Evaluating evoCas9 on endogenous genomic loci, we demonstrated a substantial improved specificity and observed no off-target sites for 4 of the 8 sgRNAs tested. Finally, we showed that following long-term expression (40 days), evoCas9 strongly limited the unspecific cleavage of a difficult-to-discriminate off-target site and fully abrogated the cleavage of two additional off-targets.

Human T cell leukemia͞lymphotropic virus type I (HTLV-I) induces adult T cell leukemia͞lymphoma (ATLL).In a small percentage of infected individuals, human T cell leukemia͞lymphotropic virus type-I (HTLV-I) causes adult T cell leukemia͞lymphoma (ATLL), an aggressive and often fatal disease (1-3). The epidemiology of ATLL suggests that cumulative genetic defects may be responsible for the acquisition of the neoplastic phenotype in a given T cell clone (4). T cell proliferation and selection following HTLV-I infection are dynamic processes that can be followed in vivo (5, 6) and in vitro (7) and generally result in the generation of clonal populations of mature CD4 ϩ , CD8 Ϫ , and CD25 ϩ ͞CD7 Ϫ T cells (8-10). In HTLV-I infection the time-dependent emergence of infected T cell clones is well documented, and ATLL results from the uncontrolled growth of a single clone.In vitro, T cell immortalization (ligand-dependent) by HTLV-I occurs within a few months of culture, whereas T cell transformation (ligand-independent) requires more time and typically results in T cell lines that display constitutive activation of the JAK͞STAT signaling pathway (11,12). In physiological conditions, activation of the JAK͞STAT pathway is triggered by cytokines through cell surface receptors (13). In the case of interferon and type I cytokines, the JAK family tyrosine kinases transduce the signal by phosphorylating the STAT proteins, which in turn dimerize and translocate to the nucleus to activate the expression of genes necessary for cell proliferation or differentiation (14). To ascertain whether the in vitro model of HTLV-I transformation has any bearing to the in vivo leukemogenesis, we investigated the JAK͞STAT activation status in uncultured ex vivo leukemic cells from 12 HTLV-I seropositive patients with ATLL.