Experimentally introduced defective endogenous proviruses are highly expressed in chickens

Abstract

We have previously described the experimental introduction of recombinant subgroup A avian leukosis viruses (ALV) with Rous-associated virus 0 long terminal repeats into the germ line of line 0 chickens and the generation of 23 transgenic lines. Two of these transgenic lines, alv6 and alvll, do not produce infectious virus. Both of these lines contain defective proviruses but do express the gag and/or env protein. We have measured viral RNA expression in tissues derived from alv6, alvll, and the parental line 0. Total RNA was prepared from 9-day embryo, 16-day embryo, 1-day chicken, and 28-day chicken tissues. Viral RNA was detected by Northern RNA transfer analysis. The results indicate that both alv6 and alvll chickens express viral RNA in all tissues tested regardless of the stage of development. No viral transcripts were detected in any line 0 (C/E; ev-negative) tissue. The levels of biologically active env glycoprotein correlates with the env RNA levels in both lines. In an in vivo interference assay, alv6, alvll, and line 0 chickens were infected with Rous-associated virus 1 and monitored for viremia, antibody against Rous-associated virus 1, and ALV-induced pathogenesis from 4 to 21 weeks. None of the 61 alv6 chickens contained detectable virus or produced antibody against subgroup A ALV. Virus and/or antibody against subgroup A ALV was detected in 34 of the 43 alvll chickens, whereas 51 of 52 line 0 birds were viremic and/or produced antibody. ALV-induced pathogenesis was observed predominantly in line 0 chickens (10 of 59), whereas very little ALV-induced pathogenesis was seen in either alv6 (1 of 62) or alvll (1 of 44) chickens. Presumably the mechanism for the increased resistance of alv6 and alvll chickens was subgroup-specific receptor interference. These results clearly demonstrate that experimentally introduced endogenous proviruses can be expressed at high levels in the avian system.

Mutations in Both the Surface and Transmembrane Envelope Glycoproteins of the RAV-2 Subgroup B Avian Sarcoma and Leukosis Virus Are Required to Escape the Antiviral Effect of a Secreted Form of the TvbS3 Receptor †

“…The presence of infectious ASLV in chickens was determined by assaying the 8-week serum samples on two different cell lines with different ASLV envelope subgroup susceptibilities: DF-1 cells (C/E) which supports the replication of ASLV(A), ASLV(B), and ASLV(C); and line alv 6 CEF (C/A) [38] which supports ASLV(B) and ASLV(C) replication. Serum samples (100 μL) were added to the cells and the cells were incubated for 9 days in media (5% serum) to allow ASLV to spread.…”

“… a Serum samples were cultured on DF-1 cells (C/E) that support the replication of ASLV(A), ASLV(B), and ASLV(C); and on line alv 6 CEFs (C/A) [38] that support ASLV(B) and ASLV(C) but not ASLV(A) replication. ELISA assays detecting ASLV capsid protein (CA) in serum were performed 8 weeks after virus challenge.…”

Soluble Forms of the Subgroup A Avian Leukosis Virus [ALV(A)] Receptor Tva Significantly Inhibit ALV(A) Infection In Vitro and In Vivo

“…Cells and animals that express retroviral envelope glycoproteins, due to a naturally occurring or genetically engineered endogenous virus, are highly resistant to retroviruses using the same receptor and have less virus-associated pathogenesis. Based on the example of the resistance of chicken lines that express endogenous subgroup E envelope glycoproteins to ALV(E) infection (38), Crittenden and colleagues demonstrated that insertion of the ALV(A) envelope gene into the germ line of chickens and its subsequent expression provided resistance to infection by ALV(A) strains by receptor interference (12,18,41,42). However, the general utility of receptor interference as an antiviral strategy may be limited for retroviruses that express cytotoxic envelope glycoproteins (17,21).…”

Single Amino Acids G196 and R198 in hr1 of Subgroup K Avian Leukosis Virus Glycoprotein Are Critical for Tva Receptor Binding

“…Most ALV-K isolates have endogenous LTR that shows considerable homology with that of endogenous virus ALV-E (Li et al, 2016;Zhao et al, 2018;Lv et al, 2019). The genomic difference between ALV-K and ALV-E was centered in SU encoded by gp85 (Figure 1) because ALV-E does not infect DF-1 (Federspiel et al, 1991); hence, the difference The supernatant was collected and diluted in gradient to determine the virus titers. Three independent experiments were performed, and data are shown as mean ± SD in triplicate from a representative experiment.…”

“…The DF-1 cell line used in this study is a continuous fibroblastic cell line derived from line 0 CEFs, which is not susceptible to endogenous virus ALV-E (Federspiel et al, 1991) but to exogenous virus ALV-K. Endogenous virus ALV-E ev-1 (GenBank: AY013303.1) and exogenous virus ALV-K GDFX0602 (GenBank: KP686143.1) show considerable homology (LTR: 98.5%; gp37: 98.9%; gp85: 87.9%) (Zhao et al, 2018), and the regions with considerable differences are primarily located in hr1 and hr2. Unlike ALV-K, ALV-E shares the Tvb-encoded tumor necrosis factor receptor with ALV-B/D as a receptor that is encoded by three alleles: tvb s1 , tvb s3 , and tvb st (Adkins et al, 1997).…”