The Functions and Mechanism of a New Oligopeptide BP9 from Avian Bursa on Antibody Responses, Immature B Cell, and Autophagy

Figure 4

BP9 induced autophagy, enhanced AMPK-ULK1 phosphorylation, and regulated the BCL-2 expression in preB cells. (a) Gene net of the differentially regulated gene involved in autophagy-related biological processes in WEHI-231 immature B cells after BP9 treatment. Purple and green circle colors indicated autophagy-related terms and regulated genes, respectively, and “+” and “−” in green circles indicated up- and downregulated genes, respectively, in BP9-treated preB cells. The bigger the purple circle area size is, the more the number of interacting genes. The bigger the green circle area size is, the more the number of interacting GO terms of these selected immune-related processes in Table S1. (b) Autophagy formation in WEHI-231 cells with 10 μg/mL BP9 treatment following immunofluorescence microscopy. Control with 10 μg/mL irrelevant peptide treatment was used as a control. The red arrow refers to autophagy in WEHI-231 cells with the swollen mitochondria. (c) LC3 protein expression with BP9 treatment in WEHI-231 cells with western blotting analysis. (d) AMPK-ULK1 phosphorylation. WEHI-231 cells were administered after BP9 treatment at experimental concentrations, total proteins were isolated, and the levels of AMPK, phosphorylated AMPK, ULK1, and phosphorylated ULK1 were analyzed by western blot using the indicated antibodies. (e) Expression levels of BCL-2. Similarly, in isolated proteins from BP9-treated WEHI-231 cells, expression levels of BCL-2 protein with BP9 treatment were assessed. In (c), (d), and (e), control with 10 μg/mL irrelevant peptide treatment was used as the irrelevant control (control) and rapamycin as the positive control. Representative western blots are demonstrated.

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