SPLUNC1 promoted cell differentiation and inhibits tumor growth.

<p>(<b>A</b>) Enforced expression of SPLUNC1 increased JNK2 and NFκB protein expression, and inhibited phosphorylation of ERK (p-Tyr-204) and Iκα (Ser-32). ΔSPLUNC1 had no effect on phosphorylation of ERK (p-Tyr-204) and Iκα (Ser-32). (<b>B</b>) SPLUNC1 also increased p27 expression and decreased the expression of CCND1, CCND2, CCND3, CCNE2, and CDK2. (<b>C, D</b>) clone formation in soft agar; tumors formed in the presence of full length SPLUNC1 or ΔSPLUNC1 were smaller versus control. (<b>E, F</b>) SPLUNC1 and ΔSPLUNC1 inhibited tumor formation in nude mice and (<b>G</b>) SPLUNC1 expression in the transplant tumor was validated by immunohistochemical analysis.</p

SPLUNC1 induced cellular apoptosis.

<p>(<b>A</b>) SPLUNC1 and the BPI domain-deleted mutant, ΔSPLUNC1, increased the expression of the proapoptotic proteins BAX, BAD caspase-3 and 8, and decreased the antiapoptotic protein Bcl-2 expression. (<b>B</b>) Enforced expression of SPLUNC1 and ΔSPLUNC1 in HNE-2 cells increased cellular apoptosis.</p

SPLUNC1 protein expression was decreased in highly differentiated nasopharyngeal carcinoma.

<p>(<b>A</b>) SPLUNC1 protein was high expressed in normal human nasopharyngeal epithelium, SPLUNC1 expression was gradually downregulated with progression from (<b>B</b>) mild to (<b>C</b>) severe atypical hyperplasia of nasopharyngeal tissues. (<b>D</b>) No positive staining for SPLUNC1 was observed in the epithelium of nasopharyngeal carcinoma.</p

SPLUNC1 expression in nasopharyngeal biopsy specimens at various stages.

<p>SPLUNC1 expression in nasopharyngeal biopsy specimens at various stages.</p

SPLUNC1 inhibited miR-141 expression and the effects of miR-141 and SPLUNC1 on PTEN expression in nasopharyngeal carcinoma cells.

<p>(<b>A</b>) SPLUNC1 expression in CNE-1 nasopharyngeal carcinoma cells was higher than that in the HNE-2 and 5–8 F cells, while the miR-141 expression in CNE-1 cells was lower than that in the HNE-2 and 5–8 F cells (P<0.05). (<b>B</b>) When SPLUNC1 was overexpressed in HNE-2 and 5–8 F cells, miR-141 expression was decreased (P<0.05). (<b>C</b>) Expression of miR-141 mimics in NPC cells decreased the expression of PTEN; expression of miR-141 inhibitor increased the expression of PTEN. (<b>D</b>) Enforced expression of SPLUNC1 in NPC cells increased PTEN expression. (<b>E</b>) miR-141 mimics inhibited PTEN expression and increased Akt phosphorylation in the presence of enforced SPLUNC1-expression in NPC 5–8 F cells. (<b>F</b>) Enforced expression of SPLUNC1 decreased Akt expression in NPC cells; and decreased the levels of phosphorylated Akt and MDM2 (<b>G</b>), but had little effect on GSK3β. SPLUNC1 inhibited the baculovirus phosphatase (BVP)-induced phosphorylation of PTEN and Akt(<b>H</b>).</p

The interaction between SPLUNC1 and LMP1.

<p>(<b>A</b>) SPLUNC1 expression was significantly decreased in NP-69 cells transfected with LMP1. (<b>B</b>) miR-141 expression was significantly increased in NP-69 cells transfected with LMP1. (<b>C</b>) PTEN expression was decreased and the Akt pathway was activated by foreign LMP1 expression. (<b>D</b>) Re-expression of SPLUNC1 in LMP1-transfected cells led to inhibition of LMP1 expression and induction of PTEN.</p

SPLUNC1 inhibited the course of EBV infection.

<p>(<b>A, B</b>) Human peripheral blood lymphocytes (LCs) were infected with green fluorescence protein (GFP)-tagged EBV. Approximately 70% of LCs showed green fluorescence after infection, but only 28% of LCs treated with the recombinant SPLUNC1 protein showed green fluorescence. (<b>C, D</b>)HNE-2 cells were co-cultured with B95-8 cells, which product EBV, for 1, 2, 3, 5, and 7 days, and B95-8 cells were then removed by complement-activated cellular cytotoxicity test. (<b>C</b>) Expression of EBER, BZLF1 and LMP1 was lower in the presence of enforced SPLUNC1 expression versus the control in HNE-2 cells. (<b>D</b>) SPLUNC1 expression in the HNE-2 cells was increased significantly 1 day after the addition of B95-8 cells to the HNE-2 culture system, and then decreased. After 3 days of co-culture with B95-8, SPLUNC1 expression decreased to its lowest level.</p