iscussion In the current study, ANXA10 was frequently overexpressed in OSCC-derived cellular lines, and down-regulation resulted in decreased cellular proliferation by inactivation of ERK. The cellcycle analysis also showed that G1 arrest occurred in ANXA10 knockdown cells through decreased CDKI expression. In addition, ANXA10 was frequently up-regulated in primary OSCCs and higher ANXA10 expression was associated with tumoral size. These results indicated that ANXA10 is linked to regulation of the cell cycle in the G1 phase and plays an important role in tumoral progression in OSCCs. Since oral carcinogenesis and other cancer types are thought to be multistep processes involving progressive disruption of epithelial cell proliferation, the mechanisms of cellular proliferation in tumoral cells are an important field of study for tumoral treatment and molecular cancer biology. Annexins, including ANXA10, have 1235481-90-9 site played important roles in tumoral development and progression. However, no direct evidence has shown that ANXA10 is required for cellular proliferation. To determine whether ANXA10 function is relevant to OSCC progression, we performed functional studies using shRNA and found that suppression of ANXA10 significantly decreased cellular proliferation as a result of inhibited MAPK signaling pathway with down-regulation of pERK. The ERK/MAPK signaling pathway is related to many cellular processes, such as proliferation, differentiation, homeostasis, and cellular survival. Activation of the ERK/MAPK signaling pathway promotes abnormal cellular growth and tumorigenesis in multiple types of tumors. Our findings indicated that ANXA10 expression was relevant to phosphorylation of ERK and activation of the ERK/MAPK ANXA10 in Oral Cancer signaling pathway. The ERK/MAPK pathway is tightly controlled by mechanisms of extracellular signals. This includes the control of intracellular Ca2+ concentrations, thereby enabling Ca2+ to serve as a second messenger. Ca2+-effector proteins can mediate cellular responses to changes in intracellular Ca2+ levels. Annexins, a family of such highly conserved Ca2+-regulated proteins, are characterized by their ability to bind to phospholipids in a Ca2+-dependent manner and most annexin functions are linked to their ability to interact with cellular membranes in a regulated manner. Based 
on these evidences combined with our results, we suggest that ANXA10 may affect intracellular Ca2+ homeostasis and directly or indirectly interacts with the activation of ERK/MAPK signaling pathways. In addition to the MAPK signaling pathway, OSCC cells with shANXA10 revealed cell-cycle arrest at the G1 phase with upregulation of p21Cip1 and p27Kip1 and down-regulation of cyclin D1 and cyclin E. The last two are also critical regulators of G1 progression and G1-S transition. Inhibiting their expression blocks G1-S transition in the cell cycle. The activities of the cyclin-CDK complexes are modulated by two types of CDKIs, Cip/Kip, which regulates cell-cycle progression. Members of the Cip/Kip family bind to cyclin-CDK complexes and inhibit their activities, which leads to G1 cell-cycle arrest. Cyclin D1, cyclin E, p21Kip1, and p27Kip1 levels are affected by multiple signaling pathways including the ERK/MAPK signaling pathway. Cyclin D1 and cyclin E are frequently upregulated in human cancers. The mechanism of activating ERK in nonadherent fibroblasts has been reported to affect cyclin D1. In other reports, activation of ERK leads t
