Ctrl siRNA + p21siRNA abc p21 β-actin Figure S1 Figure S1. Knockdown of p21 in CD4 + T cells using siRNA. Expression of p21 was analyzed by RT-PCR 36 h post transfection.  -actin was used as a house-keeping gene. P21-specific siRNA was performed using 8 nmol/ml (a), or 4nmol/ml (b) and siControl (c). This experiment is representative of repeated experiments using cells from different donors.

CD4 Annexin V -Atv + Atv (0.25 μg/ml) + Atv (0.5 μg/ml)+ Atv (1 μg/ml) Figure S2 Figure S2. Viability of CD4 + T cells following atorvastatin treatment. Representative dot plot from CD4 + T cells isolated from 1 HIV-1 seronegative individual in the absence of treatment or treated with atorvastatin (0.25 to 1  g/ml) for 4 days. Following treatment surface staining was performed. Gated cells representing percentages of apoptotic CD4 + cells as measured by flow cytometry.

p24 CD14 - Atv + Atv (0.5 μg/ml) + Atv (1 μg/ml)Negative Figure S3 Figure S3. Atorvastatin treatment limits HIV-1 replication in CD14 + cells. (A) Representative flow cytometry dot plots from CD14 + T cells in the absence or presence of atorvastatin (0.5 and 1  g/ml) after infection with X-4-tropic tropic HIV-1 isolates respectively. Data obtained by viral p24 antigen intracellular staining on day 4 post infection. These plots are representative of three repeat experiments using cells from different donors. (B) RT-PCR reflecting  -actin and p21 gene expression in monocyte-derived macrophages treated with atorvastatin (1  g/ml) for 48 h compared with untreated cells prior to total RNA isolation. A β-actin P Atorvastatin B

X4-tropic R5-tropic p24 CD4 - Atv + Atv (0.5 μg/ml) + Atv (1 μg/ml) D p24 C - Atv + Atv (0.5 μg/ml) + Atv (1 μg/ml) CD4 X4-tropic R5-tropic %CD4 + p24 + Figure S4 A B Uninfected 0

Figure S4. Atorvastatin reduces HIV-1 infection in resting CD4 + T cells. (A) Percentages of p24 suppression in CD4 + T cells that were exposed to atorvastatin (0.5-1  g/ml) for 48 h post HIV-1 infection with R5 (HIV-1 JR-CSF ) and X-4- tropic isolates. (B) Percentages of CD4 + p24 + cells in non-activated CD4 + T cells in the presence of atorvastatin (1  g/ml) 48 h after HIV-1 infection with R5 (HIV-1 JR-CSF ) and X-4-tropic (HIV-1 LAI ) isolates. Significance was tested using paired t test. (C) Representative flow cytometry dot plots from CD4 + T cells infected and then treated with  g/ml atorvastatin post infection with X-4-tropic or R5-tropic HIV-1 isolates respectively. (D) Representative flow cytometry dot plots from CD4 + T cells treated for 24 h with  g/ml atorvastatin post infection with R5 (HIV-1 JR-CSF ) and X-4-tropic (HIV-1 LAI ) isolates. CD4 + T cells from 5 HIV-1 seronegative donors were used for each viral isolate. Data are from 5 HIV-1 seronegative individuals infected with both X4-and R5-tropic viral isolates in vitro with duplicate cultures per condition. Error bars indicate mean ± SEM from duplicate cell cultures from 5 HIV-1 seronegative individuals infected 4 separate times with both X4-and R5-tropic viral isolates in vitro. Each point represents an individual.

Figure S5 siRNAp21 CD4 siControl CD4 R5-tropic HIV p24 +Atv (0.5 μg/ml) -Atv +Atv (1μg/ml) Figure S5. Atorvastatin mediated upregulation of p21 reduces HIV-1 infection in activated CD4 + T cells. Representative example of dot plots from activated CD4 + T cells infected in the presence of p21-specific or control siRNA with R5-tropic HIV-1 Isolate in the presence (0.5  g/ml or 1  g/ml) or absence of atorvastatin. Viral infection was measured by viral p24 antigen staining on CD4 + T cells using flow cytometry. These plots are representative of independent experiments performed on three different donors for each viral isolate.

- Atv + Atv (0.5 μg/ml) + Atv (1 μg/ml) + Mevalo. + Atv (0.5 μg/ml) + Mevalo. (1 mM) + Atv (1 μg/ml) + Mevalo. (1 mM) p24 CD4 X4-tropic (HIV-1 LAI ) Figure S6 Figure S6. Atorvastatin reduces infection of CD4 + T cells to HIV-1 infection and upregulates p21 through mevalonate pathway. Representative dot plots of non-activated CD4 + T cells in the presence of atorvastatin (0.5-1  g/ml) for 48 h and also in the presence or absence of L. mevalonate (1mM) post in vitro infection with X4-tropic viral isolate. Viral infection was measured by viral p24 antigen staining on CD4 + T cells using flow cytometry. These plots are representative of independent experiments performed on three different donors.