Fig. 3

Delineating the host cellular factors responsible for Ng deregulation. a Neuroblastoma cell line, SH-SY5Y cells were differentiated with RA and immunostained with MAP2-specific antibody. Green represents MAP2, and blue represents DAPI (nucleus). Ng expression increases post-differentiation of SH-SY5Y cells. Scale bar indicates 50 μm. b Differentiated (dSH-SY5Y) and undifferentiated SH-SY5Y cells were lysed and 30 μg of protein was loaded, and expression of MAP2 and Ng was measured by western blot. Tubulin was used as a loading control. c dSH-SY5Y cells were either infected with HIV-1 NLYU2 pseudotyped with VSV-G at an MOI of 1.0 for 72 h or exposed to HIV-1 or mock-infected MDM supernatants for 24 h. Expression of Ng in transduced or exposed cells was analyzed by western blot (N = 3). d Control, transduced, or exposed dSH-SY5Y cells were stained for Ng and counterstained with hematoxylin. Scale bar represents 50 μm. e Cytoplasmic and nuclear fractions of the control, transduced, or exposed dSH-SY5Y cells were examined for the presence of Ng by western blot. Lamin B and tubulin were used as nuclear and cytoplasmic markers. Nuclear and cytoplasmic Ng band intensities were normalized over lamin B and tubulin, respectively. f Cell lysates from control, transduced, or exposed dSH-SY5Y cells (30 μg) were examined for the expression of CaM, CaMKII, CREB, Syp, and Syn I by western blot. Tubulin was used as a loading control. g Relative band intensities were normalized with tubulin. Densitometry quantification of western blot data represents ±SEM of three independent observations. *p < 0.05 compared to control