FIG 1.

SPOC1 is transiently upregulated during HCMV infection. (A) HFF cells were infected with HCMV laboratory strain AD169 at an MOI of 3 and harvested at the indicated time points postinfection. Total cell extracts were prepared, separated by SDS-PAGE, and subjected to immunoblotting with mouse monoclonal antibodies p63-27 (IE1), BS 510 (pUL44), and 28-4 (MCP) and rat monoclonal SPOC1 antibody. (B) HFF cells were infected with HCMV laboratory strain AD169 at an MOI of 3. At 24 hpi, RNA was isolated with TRIzol and subsequently synthesized into cDNA via RT-PCR, and transcript levels were assessed via SYBR green PCR. The relative SPOC1 mRNA levels were calculated by normalization against the housekeeping gene RPL13A (Biomol, Hamburg, Germany). Statistical analysis was performed with Student's t test. Densitometric analysis was performed with AIDA image analyzer v.4.22 software, and SPOC1 band intensities at 24 hpi were normalized against their corresponding β-actin signals. (C and D) HFF (C) or ARPE-19 (D) cells were infected with clinical isolate TB40/E at an MOI of 3 and treated as described above for panel A. (E) Mouse embryonic fibroblasts (MEF) were infected with MCMV at an MOI of 3, and whole-cell lysates were harvested throughout the replication cycle and treated as described above for panel A. Immunoblotting was performed with the rat monoclonal SPOC1 antibody and the monoclonal mouse gB antibody. For all experiments, monoclonal antibody AC-15 (β-actin) served as a loading control.