To explore the function of AUF1 in skin cells, overexpression and RNA interference experiments were carried out. AUF1 complementary (c)DNA was obtained from OriGene Technologies, Inc. (cat. no. SC107836). AUF1 cDNA was subcloned into the pHBAD-EF1-mcs-CMV vector (Hanbio Biotechnology Co., Ltd.) for adenovirus construction. Packaging and purification of adenovirus overexpressing AUF1 (Ad-AUF1) and negative control adenovirus (Ad-NC) were carried out by Hanbio Biotechnology Co., Ltd. AUF1 cDNA was also subcloned into the pHBLV-U6-ZsGreen-Puro vector for lentivirus construction. Lentiviruses targeting AUF1 [named short hairpin (sh)AUF1-1 and shAUF1-2, respectively] and a scrambled negative control lentivirus (sh-NC) were purchased from Hanbio Biotechnology Co., Ltd. The target sequences against AUF1 are listed in Table SI . A mock-infected control group (without vector) was also constructed to demonstrate the background expression value.
HaCaT and WS1 cells were cultured in a six-well culture plate. After reaching 70% confluence, the cells were both infected with adenoviruses and lentiviruses at a multiplicity of infection of 20 with DMEM free-serum for 6 h at 37˚C in 5% CO2. After 48 h, the cells were harvested for further experiments, and the efficiency of infection was determined by reverse transcription-quantitative PCR (RT-qPCR) and western blotting.
HaCaT and WS1 cells were cultured in a six-well culture plate. After reaching 70% confluence, the cells were both infected with adenoviruses and lentiviruses at a multiplicity of infection of 20 with DMEM free-serum for 6 h at 37˚C in 5% CO2. After 48 h, the cells were harvested for further experiments, and the efficiency of infection was determined by reverse transcription-quantitative PCR (RT-qPCR) and western blotting.