Liu Cao

DNA Methylation, Its Mediators and Genome Integrity

DNA methylation regulates many cellular processes, including embryonic development, transcription, chromatin structure, X-chromosome inactivation, genomic imprinting and chromosome stability. DNA methyltransferases establish and maintain the presence of 5-methylcytosine (5mC), and ten-eleven translocation cytosine dioxygenases (TETs) oxidise 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), which can be removed by base excision repair (BER) proteins. Multiple forms of DNA methylation are recognised by methyl-CpG binding proteins (MeCPs), which play vital roles in chromatin-based transcriptional regulation, DNA repair and replication. Accordingly, defects in DNA methylation and its mediators may cause silencing of tumour suppressor genes and misregulation of multiple cell cycles, DNA repair and chromosome stability genes, and hence contribute to genome instability in various human diseases, including cancer. Thus, understanding functional genetic mutations and aberrant expression of these DNA methylation mediators is critical to deciphering the crosstalk between concurrent genetic and epigenetic alterations in specific cancer types and to the development of new therapeutic strategies.

p21-activated Kinase 6 (PAK6) Inhibits Prostate Cancer Growth via Phosphorylation of Androgen Receptor and Tumorigenic E3 Ligase Murine Double Minute-2 (Mdm2)

Background: Deregulation of AR signaling may lead to prostate cancer. Results: PAK6 promotes AR ubiquitin-mediated degradation through phosphorylation of AR and Mdm2, resulting in inhibition of prostate cancer growth in vivo. Conclusion: PAK6 suppresses prostate cancer growth via regulating AR homeostasis. Significance: Modulating PAK6 kinase activity is a therapeutic strategy for AR-positive and hormone-sensitive prostate cancer. The androgen receptor (AR) signaling pathway plays a crucial role in the development and growth of prostate malignancies. Regulation of AR homeostasis in prostate tumorigenesis has not yet been fully characterized. In this study, we demonstrate that p21-activated kinase 6 (PAK6) inhibits prostate tumorigenesis by regulating AR homeostasis. First, we demonstrated that in normal prostate epithelium, AR co-localizes with PAK6 in the cytoplasm and translocates into the nucleus in malignant prostate. Furthermore, AR phosphorylation at Ser-578 by PAK6 promotes AR-E3 ligase murine double minute-2 (Mdm2) association, causing AR degradation upon androgen stimuli. We also showed that PAK6 phosphorylates Mdm2 on Thr-158 and Ser-186, which is critical for AR ubiquitin-mediated degradation. Moreover, we found that Thr-158 collaborates with Ser-186 for AR-Mdm2 association and AR ubiquitin-mediated degradation as it facilitates PAK6-mediated AR homeostasis. PAK6 knockdown promotes prostate tumor growth in vivo. Interestingly, we found a strong inverse correlation between PAK6 and AR expression in the cytoplasm of prostate cancer cells. These observations indicate that PAK6 may be important for the maintenance of androgen-induced AR signaling homeostasis and in prostate malignancy, as well as being a possible new therapeutic target for AR-positive and hormone-sensitive prostate cancer.

PAK4 kinase-mediated SCG10 phosphorylation involved in gastric cancer metastasis

Superior cervical ganglia 10 (SCG10), as a microtubule (MT) destabilizer, maintains MT homeostasis and has a critical role in neuronal development, but its function in tumorigenesis has not been characterized. In the present study, we demonstrated that p21-activated kinase 4 (PAK4)-mediated SCG10 phosphorylation regulates MT homeostasis in metastatic gastric cancer. Our results indicate that SCG10 is a physiological substrate of PAK4, which is phosphorylated on serine 50 (Ser50) in a PAK4-dependent manner. Phosphorylated SCG10 regulated MT dynamics to promote gastric cancer cell migration and invasion in vitro and metastasis in a xenograft mouse models. Inhibiting PAK4, either by LCH-7749944 or RNA interference, resulted in the inhibition of Ser50 phosphorylation and a blockade to cell invasion, suggesting that PAK4-SCG10 signaling occurs in gastric cancer cell invasion. Moreover, we demonstrated a strong positive correlation between PAK4 and phospho-Ser50 SCG10 expression in gastric cancer samples. We also showed that high expression of SCG10 phospho-Ser50 is highly correlated to an aggressive phenotype of clinical gastric cancer. These findings revealed a novel function of SCG10 in promoting invasive potential of gastric cancer cells, suggesting that blocking PAK4-mediated SCG10 phosphorylation might be a potential therapeutic strategy for metastasis of gastric cancer.

Oncogenic PAK4 regulates Smad2/3 axis involving gastric tumorigenesis.

The alteration of p21-activated kinase 4 (PAK4) and transforming growth factor-beta (TGF-β) signaling effector Smad2/3 was detected in several types of tumors, which acts as oncogenic factor and tumor suppressor, but the relationship between these events has not been explored. Here, we demonstrate that PAK4 interacts with and modulates phosphorylation of Smad2/3 via both kinase-dependent and kinase-independent mechanisms, which attenuate Smad2/3 axis transactivation and TGF-β-mediated growth inhibition in gastric cancer cells. First, PAK4 interaction with Smad2/3, which is independent of PAK4 kinase activity, blocks TGF-β1-induced phosphorylation of Smad2 Ser465/467 or Smad3 Ser423/425 and the consequent activation. In addition, PAK4 phosphorylates Smad2 on Ser465, leading to the degradation of Smad2 through ubiquitin–proteasome-dependent pathway under hepatocyte growth factor (HGF) stimulation. Interestingly, PAK4 expression correlates negatively with phospho-Ser465/467 Smad2 but positively with phospho-Ser465 Smad2 in gastric cancer tissues. Furthermore, the expressions of HGF, phospho-Ser474 PAK4 and phospho-Ser465 Smad2 are markedly increased in gastric cancer tissues, and the expression of Smad2 is decreased in gastric cancer tissues. Our results document an oncogenic role of PAK4 in repression of Smad2/3 transactivation that involved in tumorigenesis, and suggest PAK4 as a potential therapeutic target for gastric cancer.

MDC1 functionally identified as an androgen receptor co-activator participates in suppression of prostate cancer

Mediator of DNA damage checkpoint protein 1 (MDC1) is essential for DNA damage response. However, the role of MDC1 in modulating gene transcription independently of DNA damage and the underlying mechanisms have not been fully defined. Androgen receptor (AR) is the central signaling pathway in prostate cancer (PCa) and its target genes are involved in both promotion and suppression of PCa. Here, we functionally identified MDC1 as a co-activator of AR. We demonstrate that MDC1 facilitates the association between AR and histone acetyltransferase GCN5, thereby increasing histone H3 acetylation level on cis-regulatory elements of AR target genes. MDC1 knockdown promotes PCa cells growth and migration. Moreover, depletion of MDC1 results in decreased expression of a subset of the endogenous androgen-induced target genes, including cell cycle negative regulator p21 and PCa metastasis inhibitor Vinculin, in AR positive PCa cell lines. Finally, the expression of MDC1 and p21 correlates negatively with aggressive phenotype of clinical PCa. These studies suggest that MDC1 as an epigenetic modifier regulates AR transcriptional activity and MDC1 may function as a tumor suppressor of PCa, and provide new insight into co-factor-AR-signaling pathway mechanism and a better understanding of the function of MDC1 on PCa.

Microchidia protein 2, MORC2, downregulates the cytoskeleton adapter protein, ArgBP2, via histone methylation in gastric cancer cells

ArgBP2 is an adapter protein that plays an important role in actin-dependent processes such as cell adhesion and migration. However, its function and regulation mechanisms in gastric cancer have not yet been investigated. Here, we showed the low expression of ArgBP2 mRNA level in gastric tumor samples and its repressive function in the proliferation, migration, and invasion of gastric cancer cells. Then, we cloned and identified ArgBP2 promoter and verified that MORC2 bound to the promoter. Moreover, we demonstrated that MORC2 enhanced the recruitment of EZH2, which promoted the tri-methylation of H3K27, leading to the transcriptional repression of ArgBP2. Our results might thus contribute to understanding the molecular mechanisms of ArgBP2 regulation and suggesting ArgBP2 as a potential therapeutic target for gastric cancer.

P21-activated kinase 4 involves TSH induced papillary thyroid cancer cell proliferation

Papillary thyroid cancer is a common endocrine malignancy. Although p21-activated kinase 4 (PAK4) is involved in the development of different types of tumor, its function has not been investigated in papillary thyroid cancer. Here, we identified a role for PAK4 in papillary thyroid cancer progression. Levels of PAK4 and PAK4 phosphorylated at serine 474 correlated significantly with tumor size and TNM stage. Furthermore, stable knockdown of PAK4 retarded cellular proliferation, migration, and invasion. Moreover, thyroid stimulating hormone-induced cellular proliferation in papillary thyroid cancer was found to be dependent on TSHR/cAMP/PKA/PAK4 signaling, with levels of phosphorylated PAK4 correlating positively with serum thyroid stimulating hormone and PKA Cα levels in patients with papillary thyroid cancer. These findings revealed a novel function of PAK4 in thyroid stimulating hormone-induced papillary thyroid cancer progression and suggest that PAK4 may become a promising diagnostic and therapeutic target for this disease.

miR-135a inhibits tumor metastasis and angiogenesis by targeting FAK pathway

Tumor metastasis has been the major cause of recurrence and death in patients with gastric cancer. Here, we find miR-135a has a decreased expression in the metastatic cell lines compared with its parental cell lines by analyzing microRNA array. Further results show that miR-135a is downregulated in the majority of human gastric cancer tissues and cell lines. Decreased expression of miR-135a is associated with TNM stage and poor survival. Besides, regaining miR-135a in gastric cancer cells obviously inhibits tumor growth, migration, invasion and angiogenesis by targeting focal adhesion kinase (FAK) pathway. Bioinformatics analysis and molecular experiments further prove that miR-135a is a novel downstream gene of tumor suppressor p53. Blocking FAK with its inhibitor can also enhance miR-135a expression through inducing p53. In summary, this study reveals the expression and function of miR-135a in gastric cancer and uncovers a novel regulatory mechanism of miR-135a.

BAP18 coactivates androgen receptor action and promotes prostate cancer progression

BPTF associated protein of 18 kDa (BAP18) has been reported as a component of MLL1-WDR5 complex. However, BAP18 is an uncharacterized protein. The detailed biological functions of BAP18 and underlying mechanisms have not been defined. Androgen receptor (AR), a member of transcription factor, plays an essential role in prostate cancer (PCa) and castration-resistant prostate cancer (CRPC) progression. Here, we demonstrate that BAP18 is identified as a coactivator of AR in Drosophilar experimental system and mammalian cells. BAP18 facilitates the recruitment of MLL1 subcomplex and AR to androgen-response element (ARE) of AR target genes, subsequently increasing histone H3K4 trimethylation and H4K16 acetylation. Knockdown of BAP18 attenuates cell growth and proliferation of PCa cells. Moreover, BAP18 depletion results in inhibition of xenograft tumor growth in mice even under androgen-depletion conditions. In addition, our data show that BAP18 expression in clinical PCa samples is higher than that in benign prostatic hyperplasia (BPH). Our data suggest that BAP18 as an epigenetic modifier regulates AR-induced transactivation and the function of BAP18 might be targeted in human PCa to promote tumor growth and progression to castration-resistance.

A mandatory role of nuclear PAK4-LIFR axis in breast-to-bone metastasis of ERα-positive breast cancer cells

The mechanism of estrogen receptor alpha (ERα)-positive breast cancer-associated bone metastasis is poorly understood. In this article, we report that nuclear p21-activated kinase 4 (nPAK4) is a novel repressor of ERα-mediated transactivation in a 17β-estradiol (E2)-dependent manner and promotes PAK4–ERα axis-mediated bone metastasis by targeting leukemia inhibitory factor receptor (LIFR) in ERα-positive breast cancer. An evaluation of clinical breast cancer samples revealed that nPAK4 is linked to ERα expression and appears to be associated with a poor prognosis in bone metastatic breast cancer. PAK4 bound and co-translocated with ERα from the cytoplasm to the nucleus upon stimulation with E2. nPAK4 enhanced the invasive potential of ERα-positive breast cancer cells in vitro and promoted breast cancer metastasis in vivo. Mechanistically, nPAK4 promoted the metastasis of ERα-positive breast cancer cells by targeting LIFR, a bone metastasis suppressor. Strikingly, the nuclear accumulation of PAK4 might promote aggressive phenotypes, highlighting nPAK4 as a novel predictive biomarker for ERα-positive breast cancer bone metastasis.