Limits...
Characterization of two distinct early post-entry blocks to HIV-1 in common marmoset lymphocytes

View Article: PubMed Central - PubMed

ABSTRACT

In nature, primate lentiviruses infect humans and several Old World monkeys and apes. However, to date, lentiviruses infecting New World monkeys have not been described. We studied the susceptibility of common marmoset cells to HIV-1 infection and observed the presence of post-entry blocks to the early phase of HIV-1 infection in peripheral blood lymphocytes (PBLs) and a B lymphocytic cell line (B-LCL). The blocks present in these cells are dominant and phenotypically different from each other. In PBLs, the block occurs at the level of reverse transcription, reducing the accumulation of early and late transcripts, similar to the block imposed by TRIM5α. However, we have found that marmoset TRIM5α does not block HIV-1. In contrast, the restriction factor present in B-LCLs blocks HIV-1 replication at a later step, after nuclear entry, and inhibits integration. Additionally, we have identified an HIV-1 capsid mutant, N74D, that is able to escape the restriction in the marmoset B-LCLs. Our results suggest that the factors responsible for the blocks present in marmoset PBLs and B-LCLs are different. We propose the existence of at least two new restriction factors able to block HIV-1 infection in marmoset lymphocytes.

No MeSH data available.


Related in: MedlinePlus

The post-entry block to HIV-1 in marmoset PBLs reduces the accumulation of early and late reverse transcription products.Human (hu) or marmoset (mar) PBLs were challenged with single-cycle HIV-1 viruses pseudotyped with VSV-G pretreated with DpnI for 1 h. As a control, cells were also challenged with viruses lacking an envelope glycoprotein (ΔEnv). Two hours post-infection, the cells were thoroughly washed 3 times with PBS to remove excess virus. At different time points after infection, cells were collected and total DNA isolated. The amounts of early and late RT products were quantified by real-time qPCR using TaqMan® chemistry. The results of two independent experiments are shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5120322&req=5

f4: The post-entry block to HIV-1 in marmoset PBLs reduces the accumulation of early and late reverse transcription products.Human (hu) or marmoset (mar) PBLs were challenged with single-cycle HIV-1 viruses pseudotyped with VSV-G pretreated with DpnI for 1 h. As a control, cells were also challenged with viruses lacking an envelope glycoprotein (ΔEnv). Two hours post-infection, the cells were thoroughly washed 3 times with PBS to remove excess virus. At different time points after infection, cells were collected and total DNA isolated. The amounts of early and late RT products were quantified by real-time qPCR using TaqMan® chemistry. The results of two independent experiments are shown.

Mentions: When complemented in trans with VSV-G, the single-cycle reporter viruses used in our experiments are able to enter the target cells. If the cells are permissive to infection, the virus will be able to complete all the early events in the replication cycle, including reverse transcription, nuclear entry, integration and reporter gene expression, but will not produce new viral particles. To define the step of the early HIV-1 replication cycle that is blocked in marmoset PBLs, we challenged human or marmoset PBLs with single-cycle HIV-1 green fluorescent protein (GFP) reporter viruses pseudotyped with VSV-G and analyzed the formation of early and late reverse transcription (RT) products by real-time qPCR at different times post-infection. As a control, we also infected human and marmoset PBLs with viruses lacking an envelope glycoprotein (ΔEnv). As expected, in human PBLs we detected an increase of early and late RT products over time (Fig. 4). On the contrary, in marmoset PBLs, the amounts of early and late RT products exhibited slight decreases over time (Fig. 4). These observations suggest that the factor responsible for the HIV-1 block in marmoset PBLs acts at an early step after virus entry, before or during reverse transcription. This factor may directly or indirectly disrupt the reverse transcription process itself or degrade the products of reverse transcription.


Characterization of two distinct early post-entry blocks to HIV-1 in common marmoset lymphocytes
The post-entry block to HIV-1 in marmoset PBLs reduces the accumulation of early and late reverse transcription products.Human (hu) or marmoset (mar) PBLs were challenged with single-cycle HIV-1 viruses pseudotyped with VSV-G pretreated with DpnI for 1 h. As a control, cells were also challenged with viruses lacking an envelope glycoprotein (ΔEnv). Two hours post-infection, the cells were thoroughly washed 3 times with PBS to remove excess virus. At different time points after infection, cells were collected and total DNA isolated. The amounts of early and late RT products were quantified by real-time qPCR using TaqMan® chemistry. The results of two independent experiments are shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5120322&req=5

f4: The post-entry block to HIV-1 in marmoset PBLs reduces the accumulation of early and late reverse transcription products.Human (hu) or marmoset (mar) PBLs were challenged with single-cycle HIV-1 viruses pseudotyped with VSV-G pretreated with DpnI for 1 h. As a control, cells were also challenged with viruses lacking an envelope glycoprotein (ΔEnv). Two hours post-infection, the cells were thoroughly washed 3 times with PBS to remove excess virus. At different time points after infection, cells were collected and total DNA isolated. The amounts of early and late RT products were quantified by real-time qPCR using TaqMan® chemistry. The results of two independent experiments are shown.
Mentions: When complemented in trans with VSV-G, the single-cycle reporter viruses used in our experiments are able to enter the target cells. If the cells are permissive to infection, the virus will be able to complete all the early events in the replication cycle, including reverse transcription, nuclear entry, integration and reporter gene expression, but will not produce new viral particles. To define the step of the early HIV-1 replication cycle that is blocked in marmoset PBLs, we challenged human or marmoset PBLs with single-cycle HIV-1 green fluorescent protein (GFP) reporter viruses pseudotyped with VSV-G and analyzed the formation of early and late reverse transcription (RT) products by real-time qPCR at different times post-infection. As a control, we also infected human and marmoset PBLs with viruses lacking an envelope glycoprotein (ΔEnv). As expected, in human PBLs we detected an increase of early and late RT products over time (Fig. 4). On the contrary, in marmoset PBLs, the amounts of early and late RT products exhibited slight decreases over time (Fig. 4). These observations suggest that the factor responsible for the HIV-1 block in marmoset PBLs acts at an early step after virus entry, before or during reverse transcription. This factor may directly or indirectly disrupt the reverse transcription process itself or degrade the products of reverse transcription.

View Article: PubMed Central - PubMed

ABSTRACT

In nature, primate lentiviruses infect humans and several Old World monkeys and apes. However, to date, lentiviruses infecting New World monkeys have not been described. We studied the susceptibility of common marmoset cells to HIV-1 infection and observed the presence of post-entry blocks to the early phase of HIV-1 infection in peripheral blood lymphocytes (PBLs) and a B lymphocytic cell line (B-LCL). The blocks present in these cells are dominant and phenotypically different from each other. In PBLs, the block occurs at the level of reverse transcription, reducing the accumulation of early and late transcripts, similar to the block imposed by TRIM5α. However, we have found that marmoset TRIM5α does not block HIV-1. In contrast, the restriction factor present in B-LCLs blocks HIV-1 replication at a later step, after nuclear entry, and inhibits integration. Additionally, we have identified an HIV-1 capsid mutant, N74D, that is able to escape the restriction in the marmoset B-LCLs. Our results suggest that the factors responsible for the blocks present in marmoset PBLs and B-LCLs are different. We propose the existence of at least two new restriction factors able to block HIV-1 infection in marmoset lymphocytes.

No MeSH data available.


Related in: MedlinePlus