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Fig. 4 | Retrovirology

Fig. 4

From: Uracil DNA glycosylase interacts with the p32 subunit of the replication protein A complex to modulate HIV-1 reverse transcription for optimal virus dissemination

Fig. 4

UNG2 and RPA32 expression in human cell lines and primary cells. Primary monocytes and PBMCs were isolated from blood of healthy donors. While PBMCs were activated in culture medium supplemented with PHA for 72 h and then IL-2 for 48 h, monocytes were differentiated in macrophages for 7 days in culture medium supplemented with M-CSF. a UNG2 and RPA32 protein expression. Equivalent amounts of proteins from total lysates of 293T, HeLa-CD4 and Jurkat cells, monocytes and macrophages, and non-activated and activated PBMCs (NA and A, respectively) were resolved by SDS-PAGE and analyzed by Western blotting with anti-UNG, anti-RPA32 and anti-β-actin antibodies. b, c Quantification of UNG2 and RPA32 mRNA expression. RNA was extracted and purified from 293T, HeLa-CD4, Jurkat cells, monocytes, macrophages, and non-activated and activated PBMCs, and UNG2 (b) and RPA32 (c) mRNA expression levels were then measured by quantitative RT-qPCR. Results are expressed as the percentage of UNG2 and RPA32 mRNA copies relative to those measured from 293T cell RNA extract. Data shown are means of three independent experiments (293T, HeLa and Jurkat cells) or from three independent donors for PBMCs and MDMs, performed in duplicate. Error bars represent 1 standard deviation (SD) from the mean. The inset graphs focus on mRNA expression levels in the primary cells. Negative control corresponds to 293T RNA extract processed without reverse transcriptase in the reaction mixture. ND no detection

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