Extracellular signal-regulated kinase 8 (ERK8), a recently discovered member of the mitogen-activated protein kinase protein family, has been less studied than other family members, leaving its physiologic functions mostly unknown. The biological consequences of overexpression of ERK8 in JB6 Cl41 epidermal skin cells or knockdown of ERK8 in HCT15 colorectal cancer cells was studied. Kinase assays and transient transfection experiments were performed to study the signaling pathway between ERK8 and c-Jun. We found that ERK8 is relatively highly expressed in HCT15 human colorectal cancer cells and plays an important role in the promotion and progression of colorectal cancer. ERK8 promoted neoplastic transformation, and knockdown of ERK8 in HCT15 colorectal cancer cells reduced the tumorigenic properties of these cell lines. Furthermore, a direct interaction between ERK8 and c-Jun was shown. With epidermal growth factor treatment, overexpression of ERK8 in JB6 Cl41 cells caused an increased phosphorylation of c-Jun at Ser 63 and Ser 73 , resulting in increased activator protein-1 transactivation. In contrast, knockdown of ERK8 in HCT15 colorectal cancer cells blocked c-Jun phosphorylation. The interaction between ERK8 and c-Jun seems to increase the tumorigenic properties of HCT15 colorectal cancer cells. Thus, ERK8-regulated signaling might serve as a potential therapeutic target in colorectal cancer. Cancer Res; 70(8); 3218-27. ©2010 AACR.

NEK6 (NIMA-related kinase 6) is a homologue of the Aspergillus nidulans protein NIMA (never in mitosis, gene A). We demonstrate that overexpression of NEK6 induces anchorageindependent transformation of JB6 Cl41 mouse epidermal cells. Tissue arrays and Western immunoblot analysis show that NEK6 is overexpressed in malignant tissues and several cancer cell lines. Our data also show that NEK6 interacts with STAT3, an oncogenic transcription factor, and phosphorylates STAT3 on Ser 727 , which is important for transcriptional activation. Additional studies using NEK6 mutants suggested that the phosphorylation on both Ser 206 and Thr 210 of NEK6 is critical for STAT3 phosphorylation and anchorage-independent transformation of mouse epidermal cells. Notably, knockdown of NEK6 decreased colony formation and STAT3 Ser 727 phosphorylation. Based on our findings, the most likely mechanism that can account for this biological effect involves the activation of STAT3 through the phosphorylation on Ser 727 . Because of the critical role that STAT3 plays in mediating oncogenesis, the stimulatory effects of NEK6 on STAT3 and cell transformation suggest that this family of serine/threonine kinases might represent a novel chemotherapeutic target.

Various types of post-translational modifications of the histone tails have been revealed, but a few modifications have been found within the histone core sequences. Histone core posttranslational modifications have the potential to modulate nucleosome structure and DNA accessibility. Here, we studied the histone H2B core domain and found that phosphorylation of H2B serine 32 occurs in normal cycling and mitogen-stimulated cells. Notably, this phosphorylation is elevated in skin cancer cell lines and tissues compared with normal counterparts. The JB6 Cl41 mouse skin epidermal cell line is a well established model for tumor promoter-induced cell transformation and was used to study the function of H2B during EGF-induced carcinogenesis. Remarkably, cells overexpressing a nonphosphorylatable H2BS32A mutant exhibited suppressed growth and EGFinduced cell transformation, possibly because of decreased activation of activator protein-1, compared with control cells overexpressing wild type H2B. We identified ribosomal S6 kinase 2 (RSK2) as the kinase responsible for H2BS32 phosphorylation. Serum-starved JB6 cells contain very little endogenous H2BS32 phosphorylation, and EGF treatment induced this phosphorylation. The phosphorylation was attenuated in RSK2 knock-out MEFs and RSK2 knockdown JB6 cells. Taken together, our results demonstrate a novel role for H2B phosphorylation in cell transformation and show that H2BS32 phosphorylation is critical for controlling activator protein-1 activity, which is a major driver in cell transformation.The eukaryote genome is packaged into the highly organized chromatin. The fundamental repeating unit of chromatin is the nucleosome, which consists of 146 bp of DNA wrapped approximately two turns around a core histone octamer (two copies each of histone H2A, H2B, H3, and H4) (1). Generally speaking, chromatin is inaccessible to the outside environment because of the strong DNA/histone(s) and histone(s)/histone(s) interactions. Thus, the chromatin structure must be remodeled in different ways to facilitate any process that requires access to the DNA (2).The chromatin structure is regulated by four main processes: 1) post-translational modifications of histone variants; 2) incorporation (or replacement) of histone variants; 3) DNA methylation; and 4) ATP-dependent chromatin remodeling (3). The post-translational modifications (e.g. acetylation, methylation, phosphorylation, ubiquitylation, sumoylation, or proline isomerization) of histone variants have drawn much attention and are a primary focus in the field of epigenetics (4). Some studies suggest that these modifications are performed by modifiers with multiple chromatin-binding domains to engage histone tails (5). Evidence also indicates that combinations of posttranslational histone modifications cooperate to increase the specificity of the signal to be elicited or translated to distinct biological events (6).Histone tails protrude from their own nucleosome, and they are able to make contact with adjacent nucleosomes. Thus, modifica...