Differential Evolution of Antiretroviral Restriction Factors in Pteropid Bats as Revealed by APOBEC3 Gene Complexity

Hayward, Joshua A; Tachedjian, Mary; Cui, Jie; Cheng, Adam Z; Johnson, Adam; Baker, Michelle L; Harris, Reuben S; Wang, Lin-Fa; Tachedjian, Gilda

Approval for the use of bat tissues was granted by the Commonwealth Scientific and Industrial Research Organization's Australian Animal Health Laboratory Animal Ethics Committee (Protocol AEC1281). Transcriptome data sets of P. alecto were generated from pooled total RNA from mitogen-stimulated spleen, lymph node, white blood cells, and unstimulated bone-marrow and thymus from one pregnant female and one adult male, and the unstimulated thymus of a juvenile male as described previously (Papenfuss et al. 2012 Ensembl: ENST00000348946] . The human A3 transcripts were translated into protein sequences using the CLC Genomics Workbench 8.0 (https://www.qiagenbioinformatics.com/; last accessed April 3, 2018). To identify transcripts of interest the tBLASTn function of CLC Genomics Workbench was used with the following parameters: BLOSUM62 matrix, word size ¼ 3, E-values < 1 Â 10 À12 , gap costs of existence 11, extension 1, with no filtering of regions of low complexity. Transcripts were confirmed as A3 homologues by initial identification of, in their translated protein sequences, the presence of the conserved Z-domain, Hx 1 Ex 21-27 S/TWSPCx 2-4 C (where "x" indicates a nonconserved position [Conticello et al. 2005; LaRue et al. 2009 ]), and subsequent identification of the A3 subtype-specific conserved motifs described in detail in LaRue et al. (2008, 2009 ).