MYG1 drives glycolysis and colorectal cancer development through nuclear-mitochondrial collaboration.

Metabolic remodeling is a strategy for tumor survival under stress. However, the molecular mechanisms during the metabolic remodeling of colorectal cancer (CRC) remain unclear. Melanocyte proliferating gene 1 (MYG1) is a 3'-5' RNA exonuclease and plays a key role in mitochondrial functions. Here, we uncover that MYG1 expression is upregulated in CRC progression and highly expressed MYG1 promotes glycolysis and CRC progression independent of its exonuclease activity. Mechanistically, nuclear MYG1 recruits HSP90/GSK3ß complex to promote PKM2 phosphorylation, increasing its stability. PKM2 transcriptionally activates MYC and promotes MYC-medicated glycolysis. Conversely, c-Myc also transcriptionally upregulates MYG1, driving the progression of CRC. Meanwhile, mitochondrial MYG1 on the one hand inhibits oxidative phosphorylation (OXPHOS), and on the other hand blocks the release of Cyt c from mitochondria and inhibits cell apoptosis. Clinically, patients with KRAS mutation show high expression of MYG1, indicating a high level of glycolysis and a poor prognosis. Targeting MYG1 may disturb metabolic balance of CRC and serve as a potential target for the diagnosis and treatment of CRC.

EphB6 deficiency in intestinal neurons promotes tumor growth in colorectal cancer by neurotransmitter GABA signaling.

EphB6 belongs to the receptor tyrosine kinase, whose low expression is associated with shorter survival of colorectal cancer (CRC) patients. But the role and mechanism of EphB6 in the progression of CRC need further study. In addition, EphB6 was mainly expressed in intestinal neurons. But how EphB6 is involved in functions of intestinal neurons has not been known. In our study, we constructed a mouse xenograft model of CRC by injecting CMT93 cells into the rectum of EphB6-deficient mice. We found that the deletion of EphB6 in mice promoted tumor growth of CMT93 cells in a xenograft model of CRC, which was independent of changes in the gut microbiota. Interestingly, inhibition of intestinal neurons by injecting botulinum toxin A into rectum of EphB6-deficient mice could eliminate the promotive effect of EphB6 deficiency on tumor growth in the xenograft model of CRC. Mechanically, the deletion of EphB6 in mice promoted the tumor growth in CRC by increasing GABA in the tumor microenvironment. Furthermore, EphB6 deficiency in mice increased the expression of synaptosomal-associated protein 25 in the intestinal myenteric plexus, which mediated the release of GABA. Our study concluded that EphB6 knockout in mice promotes tumor growth of CMT93 cells in a xenograft model of CRC by modulating GABA release. Our study found a new regulating mechanism of EphB6 on the tumor progression in CRC that is dependent on intestinal neurons.

PLOD2 promotes colorectal cancer progression by stabilizing USP15 to activate the AKT/mTOR signaling pathway.

Procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) has been reported as an oncogenic gene, affecting various malignant tumors, including endometrial carcinoma, osteosarcoma, and gastric cancer. These effects are mostly due to the enhanced deposition of collagen precursors. However, more studies need to be conducted on how its lysyl hydroxylase function affects cancers like colorectal carcinoma (CRC). Our present results showed that PLOD2 expression was elevated in CRC, and its higher expression was associated with poorer survival. Overexpression of PLOD2 also facilitated CRC proliferation, invasion, and metastasis in vitro and in vivo. In addition, PLOD2 interacted with USP15 by stabilizing it in the cytoplasm and then activated the phosphorylation of AKT/mTOR, thereby promoting CRC progression. Meanwhile, minoxidil was demonstrated to downregulate the expression of PLOD2 and suppress USP15, and the phosphorylation of AKT/mTOR. Our study reveals that PLOD2 plays an oncogenic role in colorectal carcinoma, upregulating USP15 and subsequently activating the AKT/mTOR pathway.

LARP6 suppresses colorectal cancer progression through ZNF267/SGMS2-mediated imbalance of sphingomyelin synthesis.

BACKGROUND: With increasing incidence and mortality, colorectal cancer (CRC) seriously endangers human health. LARP6, a member of La-related protein (LARP) family, is a RNA binding protein and probably associates with CRC progression, but its specific roles and mechanisms in CRC still remain unknown. METHOD: Quantitative real-time PCR (qPCR), western blot, and immunohistochemistry were employed to examine LARP6 expression in CRC tissues. Using the stable LARP6 overexpression or interference CRC cell lines, the effect of LARP6 on CRC progression were evaluated. High-throughput RNA immunoprecipitation sequencing (RIP-seq) and a series of relevant experiments were conducted to explain how LARP6 functions. SPSS software was used for statistical analysis. RESULT: In this study, we found that LARP6 expression is downregulated in CRC and correlates with patients' overall survival and relapse-free survival. Furthermore, altered LARP6 expression influences CRC cells invasion and metastasis. Mechanically, we discovered that LARP6 bind ZNF267 mRNA and regulated its stability and translation. LARP6 inhibited expression of SGMS2, a downstream target of ZNF267, resulting in ceramide and sphingomyelin imbalance in CRC cells. Interestingly, LARP6 also enhances autophagy activity of CRC cells, and the effect was at least partially determined by the inhibition of SGMS2-mediated sphingomyelin synthesis. CONCLUSION: Our study showed how LARP6/ZNF267/SGMS2 axis influence CRC progression, which contributes to further understanding of the molecular mechanisms underlying CRC development.

LINC00839 promotes colorectal cancer progression by recruiting RUVBL1/Tip60 complexes to activate NRF1.

The long noncoding RNA LINC00839 has been shown to be involved in the progression of some cancer types, such as bladder cancer, prostate cancer, breast cancer, and neuroblastoma. However, if LINC00839 has roles in colorectal cancer (CRC), it has not been elucidated so far. Here, we focus on the biological role and involved mechanisms of LINC00839 in CRC. We show that LINC00839 is selectively upregulated in CRC and locates to the nucleus. High expression of LINC00839 is associated with poor outcomes in CRC patients. Functional experiments show that LINC00839 promotes CRC proliferation, invasion, and metastasis in vitro and in vivo. Mechanistically, LINC00839 recruits Ruvb1 to the Tip60 complex and increases its acetylase activity. LINC00839 guides the complex to the NRF1 promoter and promotes acetylation of lysines 5 and 8 of histones H4, thereby upregulating the expression of NRF1. Subsequently, NRF1 activates mitochondrial metabolism and biogenesis, thereby promoting CRC progression. In summary, our study reports on a mechanism by which LINC00839 positively regulates NRF1, thus promoting mitochondrial metabolism and biogenesis, as well as CRC progression.

RSL1D1 promotes the progression of colorectal cancer through RAN-mediated autophagy suppression.

RSL1D1 (ribosomal L1 domain containing 1), a member of the universal ribosomal protein uL1 family, was suggested to be a new candidate target for colorectal cancer (CRC). However, the role of RSL1D1 in cancer, including CRC, remains largely elusive. Here, we demonstrated that RSL1D1 expression was significantly elevated in tumors from CRC patients and that high expression of RSL1D1 was correlated with poorer survival of CRC patients. Functionally, RSL1D1 promoted the proliferation, invasion, and metastasis of CRC cells by suppressing autophagy. Interestingly, RSL1D1 interacted with RAN and inhibited its deacetylation by competitively binding with Sirt7. By affecting the acetylation of RAN, RSL1D1 inhibited the accumulation of nuclear STAT3 and the STAT3-regulated autophagic program. Taken together, our study uncovered the key role of the RSL1D1/RAN/STAT3 regulatory axis in autophagy and tumor progression in CRC, providing a new candidate target for CRC treatment.

MIER3 induces epithelial-mesenchymal transition and promotes breast cancer cell aggressiveness via forming a co-repressor complex with HDAC1/HDAC2/Snail.

Breast cancer is one of the most frequently diagnosed cancers and the leading cause of cancer death in women. MIER3 (Mesoderm induction early response 1, family member3) is considered as a potential oncogene for breast cancer. However, the role of MIER3 in breast cancer remain largely unknown. The expression of MIER3 was detected and the relationship between its expression and clinicopathological characteristics was also analyzed. The effect of MIER3 on proliferation and migration of breast cancer cells was detected in vitro and in vivo. Western blot, IF, and Co-IP were employed to detect the relationship between MIER3, HDAC1, HDAC2, and Snail. ChIP assay was performed to determine the binding of MIER3/HDAC1/HDAC2/Snail complex to the promoter of E-cadherin. In this study, we found that MIER3 was upregulated in breast cancer tissue and closely associated with poor prognosis of patients. MIER3 could promote the proliferation, migration, and epithelial-mesenchymal transition (EMT) of breast cancer cells. Further studies showed that MIER3 interacted with HDAC1/HDAC2 and Snail to form a repressive complex which could bind to E-cadherin promoter and was related to its deacetylation. Our study concluded that MIER3 was involved in forming a co-repressor complex with HDAC1/HDAC2/Snail to promote EMT by silencing E-cadherin.

CSRP2 suppresses colorectal cancer progression via p130Cas/Rac1 axis-meditated ERK, PAK, and HIPPO signaling pathways.

Metastasis is a major cause of death in patients with colorectal cancer (CRC). Cysteine-rich protein 2 (CSRP2) has been recently implicated in the progression and metastasis of a variety of cancers. However, the biological functions and underlying mechanisms of CSRP2 in the regulation of CRC progression are largely unknown. Methods: Immunohistochemistry, quantitative real-time polymerase chain reaction (qPCR) and Western blotting (WB) were used to detect the expression of CSRP2 in CRC tissues and paracancerous tissues. CSRP2 function in CRC was determined by a series of functional tests in vivo and in vitro. WB and immunofluorescence were used to determine the relation between CSRP2 and epithelial-mesenchymal transition (EMT). Co-immunoprecipitation and scanning electron microscopy were used to study the molecular mechanism of CSRP2 in CRC. Results: The CSRP2 expression level in CRC tissues was lower than in adjacent normal tissues and indicated poor prognosis in CRC patients. Functionally, CSRP2 could suppress the proliferation, migration, and invasion of CRC cells in vitro and inhibit CRC tumorigenesis and metastasis in vivo. Mechanistic investigations revealed a physical interaction between CSRP2 and p130Cas. CSRP2 could inhibit the activation of Rac1 by preventing the phosphorylation of p130Cas, thus activating the Hippo signaling pathway, and simultaneously inhibiting the ERK and PAK/LIMK/cortactin signaling pathways, thereby inhibiting the EMT and metastasis of CRC. Rescue experiments showed that blocking the p130Cas and Rac1 activation could inhibit EMT induced by CSRP2 silencing. Conclusion: Our results suggest that the CSRP2/p130Cas/Rac1 axis can inhibit CRC aggressiveness and metastasis through the Hippo, ERK, and PAK signaling pathways. Therefore, CSRP2 may be a potential therapeutic target for CRC.

Prognostic value of delta-like protein 3 combined with thyroid transcription factor-1 in small-cell lung cancer.

Rovalpituzumab tesirine is a promising delta-like protein 3 (DLL3)-targeted antibody-drug conjugate for the treatment of small-cell lung cancer (SCLC). Thyroid transcription factor-1 (TTF-1) and DLL3 protein are associated with SCLC, and may be used to identify patients, who respond to the DLL3-targeted therapy. However, little is known about the expression pattern of the DLL3 protein, and the prognostic value of DLL3 and TTF-1 for SCLC. A total of 335 patients with SCLC were identified, including 11 patients with paired biopsy of primary site and lobectomy specimens, and 37 patients with paired specimens of primary and metastatic site. The DLL3 expression levels of individuals were evaluated using the anti-DLL3 antibody. No differences in DLL3 expression levels were observed in paired biopsy and lobectomy specimens (P=0.774), and paired primary and metastatic sites (P=0.472). SCLC cases with high DLL3 expression levels were more frequent in male patients (P=0.041), smokers (P=0.023) and patients with positive TTF-1 expression (P=0.006) compared with DLL3-low SCLC. DLL3-high SCLC exhibited worse overall survival compared with DLL3-low SCLC (log-rank test, P=0.007). Patients with TTF-1+ SCLC experienced a significantly worse overall survival compared with patients with TTF-1- SCLC (P<0.001). DLL3-low/TTF-1- was defined as a distinct molecular subgroup of SCLC with optimal prognosis (P<0.001). DLL3-low/TTF-1- was an independent prognostic marker for SCLC (P=0.001). In conclusion, the present study, to the best of our knowledge, provided novel evidence for SCLC intratumoral and intertumoral homogeneity with the identification of DLL3 protein levels. Therefore, it is reliable to use biopsy specimens to evaluate DLL3 expression levels for identification of patients who may benefit from DLL3-targeted therapy. In addition, DLL3 and TTF-1 are two protein markers with potential clinical value in risk stratification for patients with SCLC.

Complete mitochodrial genome of Parasinilabeo assimilis (Cypriniformes, Cyprinidae).

The complete mitochondrial genome of Parasinilabeo assimilis has been determined in this study. The gene composition, arrangement and transcriptional orientation in P. assimilis mitogenome were identical to most vertebrates. The complete mitogenome of P. assimilis was 16,602 bp in size with 13 protein coding genes, 22 tRNA genes and a control region. Two start codon patterns and three stop codon patterns were found in protein-coding genes. Only the tRNA-Ser2 could not fold into a typical clover-leaf secondary structure due to the lack of the dihydrouridine arm. Sequence alignment results suggest that the complete mitogenome of P. assimilis is an efficient tool to study molecular phylogenetics, biogeography and adaptive evolution of this lineage.

[Detection of KRAS,NRAS and BRAF gene mutations in colorectal carcinoma].

OBJECTIVE: To investigate mutations frequencies of KRAS,NRAS and BRAF genes in colorectal carcinoma. METHODS: Tissue specimens from 200 colorectal cancer patients at diagnosis were collected and subject to KRAS,NRAS and BRAF mutation analyses by PCR-based direct DNA sequencing targeting exons 2, 3 and 4 of KRAS gene, exons 2, 3 and 4 of NRAS gene and exon 15 of BRAF gene. RESULTS: Activating mutations were detected in KRAS (44%, 88/200), NRAS (2%, 4/200) and BRAF (5%, 10/200) in this study cohort.Among KRAS mutations, 64.8% (57/88) occurred in codon 12 and 12.5% (11/88) occurred in codon 13. KRAS gene mutation in exon 3 mainly involved codons 59 and 61. KRAS gene mutation in exon 4 mainly involved codons 117 and 146. CONCLUSIONS: Mutations at exon 2 of KRAS gene have the highest frequency in colorectal carcinoma. Expanding the detection sites of KRAS gene combined with NRAS and BRAF genes may help to identify patients who will most likely benefit from targeted therapies.

A signature for induced pluripotent stem cell-associated genes in colorectal cancer.

Genes associated with induced pluripotent stem cells (iPS genes) are several pivotal transcriptional factors, which are used to induce pluripotent stem cells from some adult somatic cells. The roles of these iPS genes and especially the signature for these iPS genes in colorectal cancer (CRC) are still unclear. Overexpressed Oct4 and Lin28 but down-regulated Nanog were found in tumor tissues compared with that in their paired normal counterparts of CRC patients. Interestingly, we found that Oct4, Lin28 and Nanog were highly overexpressed in some patients. And the signature for iPS genes was correlated with tumor site (P = 0.012), lymph node status (P = 0.033), Dukes classification (P = 0.033) of CRC patients. Moreover, an independent public expression profiling data showed signature for the four iPS genes could successfully be used to predict the survival of CRC patients with Dukes stages B and C. Immunofluorescent staining of fresh CRC tissues from patients showed that strong co-expressions of Oct4 and Nanog proteins or Sox2 and Lin28 were present in some CRC cells. Then, CRC cell subclone with four iPS genes overexpression were establish by a mixed retroviral system. We found that iPS genes promote sphere-formation, proliferation, colony formation, migration of human CRC cells in vitro and tumor growth in vivo. Our study first shows the clinical significance of iPS signature in CRC patients.

An epigenetic role for PRL-3 as a regulator of H3K9 methylation in colorectal cancer.

OBJECTIVE: This study investigated the epigenetic role of PRL-3, a key metastasis gene in colorectal cancer (CRC), as a regulator of histone demethylation and the functions of Jumonji domain-containing protein 1B (JMJD1B) and JMJD2B in the progression of CRC. METHODS: PRL-3-associated proteins were analysed using functional distribution and category enrichment analysis. Western blotting and immunofluorescence were used to detect nuclear PRL-3. The relationship between PRL-3 and JMJD1B or JMJD2B and the roles of JMJD1B, JMJD2B and PRL-3 in histone demethylation were determined after these proteins were knocked down using RNA interference. Case-control studies on JMJD1B and JMJD2B in patients with CRC were performed using immunohistochemical analysis. The in vitro functional effects of JMJD2B and JMJD1B were examined further. RESULTS: JMJD1B and JMJD2B, two histone demethylases, were enriched among PRL-3-associated proteins. Nuclear PRL-3 was observed in CRC cells and clinical samples of CRC. The expression of nuclear PRL-3 was increased in patients with CRC at more advanced Dukes' stages. PRL-3 was involved in the regulation of histone methylation by affecting the activities of JMJD1B and JMJD2B. A low expression of the JMJD1B protein was positively correlated with the lymph node status (p=0.032), Dukes' classification (p=0.008) and TNM staging (p=0.022) of patients with CRC. A high expression of JMJD2B was positively correlated with the lymph node status (p=0.03), Dukes' classification (p=0.036) and tumour invasion (p=0.003) of patients with CRC. A loss-of-function analysis confirmed that JMJD2B promoted the proliferation, colony formation and migration of human CRC cells. CONCLUSION: Our data reveal a new role for PRL-3 as a key regulator of histone demethylation. JMJD1B seems to be a candidate tumour suppressor and JMJD2B seems to be a potential oncoprotein in the development and progression of CRC.

Snail as a key regulator of PRL-3 gene in colorectal cancer.

The regulators of a key metastasis gene PRL-3 in colorectal cancer (CRC) are still largely unknown. We found three potential binding sites of Snail, a key transcriptional factor involved in the epithelial-mesenchymal transition (EMT), in the region of PRL-3 promoter (located at -642 to -383). Moreover, our results showed that one of the Snail binding sites (located at -624 to -619) was the key element to maintain promoter activity of human PRL-3 gene. The transcriptional activity of PRL-3 promoter was abolished after the Snail binding site (located at -624 to -619) was mutated. Both promoter activity and protein expression of PRL-3 in CRC cell lines could be regulated by Snail. In clinical samples of CRC and metastatic lymph node of CRC, expression of PRL-3 protein was correlated with expression of Snail protein. Functional studies using gene over-expression and knockdown methods indicated that Snail promoted proliferation, cell adhesion and migration of human CRC cells. In SW480 cells with PRL-3 stable knockdown, cell proliferation increased after Snail was up-regulated. Our data first reveal transcriptional factor Snail as a key regulator of PRL-3 in CRC. The link between Snail and PRL-3 suggests a new potential mechanism of Snail contributing to progression and metastasis of CRC.

Stathmin, a new target of PRL-3 identified by proteomic methods, plays a key role in progression and metastasis of colorectal cancer.

To better understand the role of PRL-3 in progression and metastasis of colorectal cancer (CRC), we searched for PRL-3 associated proteins using proteomic methods. We identified 39 PRL-3 associated proteins based on proteomic strategy. Stathmin, a key oncoprotein, was proved to be a new PRL-3 associated protein. Notably, co-immunoprecipitation assays in both endogenous CRC cell lines and CRC tissues indicated that PRL-3 could interact with stathmin. And, both stathmin and PRL-3 contributed to microtubule (MT) destabilization of CRC cells. Moreover, gain-of-function and loss-of-function analyses revealed that stathmin promoted proliferation, cell adhesion, and migration of human CRC cells. Immunohistochemical analysis of 149 colorectal tumor samples showed that overexpression of stathmin was strongly correlated with tumor differentiation (P = 0.035), tumor invasion (P = 0.024), lymph node status (P < 0.001), Dukes classification (P < 0.001), and TNM staging (P < 0.001) of CRC patients. Univariate and multivariate survival analyses further supported that overexpression of stathmin protein was a potential independent poor prognostic factor for CRC. Our results reveal many PRL-3 associated proteins for the first time. The oncoprotein stathmin plays a key role in CRC as a new target of PRL-3. Interaction between PRL-3 and stathmin leads to MT destabilization of CRC cells, which contributes to progression and metastasis of CRC.