Nrf1 Is Endowed with a Dominant Tumor-Repressing Effect onto the Wnt/β-Catenin-Dependent and Wnt/β-Catenin-Independent Signaling Networks in the Human Liver Cancer

Our previous work revealed that Nrf1α exerts a tumor-repressing effect because its genomic loss (to yield Nrf1α-/-) results in oncogenic activation of Nrf2 and target genes. Interestingly, β-catenin is concurrently activated by loss of Nrf1α in a way similar to β-catenin-driven liver tumor. However, a presumable relationship between Nrf1 and β-catenin is not yet established. Here, we demonstrate that Nrf1 enhanced ubiquitination of β-catenin for targeting proteasomal degradation. Conversely, knockdown of Nrf1 by its short hairpin RNA (shNrf1) caused accumulation of β-catenin so as to translocate the nucleus, allowing activation of a subset of Wnt/β-catenin signaling responsive genes, which leads to the epithelial-mesenchymal transition (EMT) and related cellular processes. Such silencing of Nrf1 resulted in malgrowth of human hepatocellular carcinoma, along with malignant invasion and metastasis to the lung and liver in xenograft model mice. Further transcriptomic sequencing unraveled significant differences in the expression of both Wnt/β-catenin-dependent and Wnt/β-catenin-independent responsive genes implicated in the cell process, shape, and behavior of the shNrf1-expressing tumor. Notably, we identified that β-catenin is not a target gene of Nrf1, but this CNC-bZIP factor contributes to differential or opposing expression of other critical genes, such as CDH1, Wnt5A, Wnt11A, FZD10, LEF1, TCF4, SMAD4, MMP9, PTEN, PI3K, JUN, and p53, each of which depends on the positioning of distinct cis-regulatory sequences (e.g., ARE and/or AP-1 binding sites) in the gene promoter contexts. In addition, altered expression profiles of some Wnt/β-catenin signaling proteins were context dependent, as accompanied by decreased abundances of Nrf1 in the clinic human hepatomas with distinct differentiation. Together, these results corroborate the rationale that Nrf1 acts as a bona fide dominant tumor repressor, by its intrinsic inhibition of Wnt/β-catenin signaling and relevant independent networks in cancer development and malignant progression.

HTLV-1 bZIP factor: the key viral gene for pathogenesis

Human T cell leukemia virus type 1 (HTLV-1) causes adult T-cell leukemia-lymphoma (ATL) and inflammatory diseases. The HTLV-1 bZIP factor (HBZ) gene is constantly expressed in HTLV-1 infected cells and ATL cells. HBZ protein suppresses transcription of the tax gene through blocking the LTR recruitment of not only ATF/CREB factors but also CBP/p300. HBZ promotes transcription of Foxp3, CCR4, and T-cell immunoreceptor with Ig and ITIM domains (TIGIT). Thus, HBZ is critical for the immunophenotype of infected cells and ATL cells. HBZ also functions in its RNA form. HBZ RNA suppresses apoptosis and promotes proliferation of T cells. Since HBZ RNA is not recognized by cytotoxic T cells, HTLV-1 has a clever strategy for avoiding immune detection. HBZ plays central roles in maintaining infected T cells in vivo and determining their immunophenotype.

Nrf1 is endowed with a dominant tumor-repressing effect onto the Wnt/β-Catenin-dependent and -independent signaling networks in the human liver cancer

ABSTRACTOur previous work revealed that Nrf1α exerts a tumor-repressing effect because its genomic loss (to yield Nrf1α−/−) results in oncogenic activation of Nrf2 and target genes. Interestingly, β-catenin is concurrently activated by loss of Nrf1α in a way similar to β-catenin-driven liver tumor. However, a presumable relationship between Nrf1 and β-catenin is not as yet established. Here, we demonstrate that Nrf1 enhanced ubiquitination of β-catenin for targeting to proteasomal degradation. Conversely, knockdown of Nrf1 by its short-hairpin RNA (shNrf1) caused accumulation of β-catenin so as to translocate the nucleus, allowing activation of a subset of Wnt–β-catenin signaling responsive genes, which leads to the epithelial-mesenchymal transition (EMT) and related cellular processes. Such silencing of Nrf1 resulted in malgrowth of human hepatocellular carcinoma, along with malignant invasion and metastasis to the lung and liver in xenograft model mice. Further transcriptomic sequencing unraveled significant differences in the expression of both Wnt/β-catenin-dependent and -independent responsive genes implicated in the cell process, shape and behavior of the shNrf1-expressing tumor. Notably, we identified that β-catenin is not a target gene of Nrf1, but this CNC-bZIP factor contributes to differential or opposing expression of other critical genes, such as CDH1, Wnt5A, Wnt11A, FZD10, LEF, TCF4, SMAD4, MMP9, PTEN, PI3K, JUN and p53, each of which depends on the positioning of distinct cis-regulatory sequences (e.g., ARE and/or AP-1 binding sites) in the gene promoter contexts. In addition, altered expression profiles of some Wnt–β-catenin signaling proteins were context-dependent, as accompanied by decreased abundances of Nrf1 in the clinic human hepatomas with distinct differentiation. Together, these results corroborate the rationale that Nrf1 acts as a bona fide dominant tumor-repressor, by its intrinsic inhibition of Wnt–β-catenin signaling and relevant independent networks in cancer development and malignant progression.

AVALIAÇÃO DA PROTEÍNA HBZ NO DESENVOLVIMENTO DA LEUCEMIA/LINFOMA DE CÉLULAS T DO ADULTO: UMA REVISÃO SISTEMÁTICA

A proteína HTLV-1 bzip factor (HBZ) é produzida pelas células infectadas pelo HTLV-1 e sua expressão se dá de forma constante em células de indivíduos com Leucemia/Linfoma de Células T do Adulto (ATLL). Essa patologia é grave e, apesar de existir tratamentos que reduzam os sintomas e aumentem a sobrevida do paciente, ainda não existe cura conhecida para a infecção pelo HTLV-1. A HBZ foi descrita como uma das principais proteínas virais que induzem proliferação e imortalização celular da célula infectada pelo HTLV-1. Como a HBZ é sempre expressa em células de pacientes com ATLL, é importante discutir os mecanismos de atuação dessa proteína para auxiliar estudos futuros sobre tratamento e prevenção de doenças provocadas pelo HTLV-1. Portanto, esse trabalho possui como objetivo principal investigar as vias de atuação dessa proteína no desenvolvimento da ATLL. Essa revisão sistemática foi realizada a partir de artigos encontrados na plataforma de busca do PubMed. Foram encontrados 61 artigos, destes 20 foram incluídos e seus dados foram coletados. A partir dos dados obtidos, é possível concluir que proteína HBZ é importante no desenvolvimento de ATLL ao contribuir na proliferação, imortalização e sobrevivência das células infectadas. Porém ainda não está claro se a HBZ participa do mecanismo inicial de desenvolvimento da ATLL.

A Naturally-Occurring Dominant-Negative Inhibitor of Keap1 Competitively against Its Negative Regulation of Nrf2

Transcription factor Nrf2 (nuclear factor-erythroid 2-related factor 2) is a master regulator of antioxidant and/or electrophile response elements (AREs/EpREs)-driven genes involved in homeostasis, detoxification, and adaptation to various stresses. The cytoprotective activity of Nrf2, though being oppositely involved in both cancer prevention and progression, is critically controlled by Keap1 (Kelch-like ECH-associated protein 1), which is an adaptor subunit of Cullin 3-based E3 ubiquitin ligase and also is a key sensor for oxidative and electrophilic stresses. Here, we first report a novel naturally-occurring mutant of Keap1, designated Keap1ΔC, which lacks most of its C-terminal Nrf2-interacting domain essential for inhibition of the cap’n’collar (CNC) basic-region leucine zipper (bZIP) factor. This mutant Keap1ΔC is yielded by translation from an alternatively mRNA-spliced variant lacking the fourth and fifth exons, but their coding sequences are retained in the wild-type Keap1 locus (with no genomic deletions). Although this variant was found primarily in the human highly-metastatic hepatoma (MHCC97H) cells, it was widely expressed at very lower levels in all other cell lines examined. Such Keap1ΔC retains no or less ability to inhibit Nrf2, so that it functions as a dominant-negative competitor of Keap1 against its inhibition of Nrf2 due to its antagonist effect on Keap1-mediated turnover of Nrf2 protein.

Topovectorial mechanisms control the juxtamembrane proteolytic processing of Nrf1 to remove its N-terminal polypeptides during maturation of the CNC-bZIP factor

ABSTRACTThe topobiological behaviour of Nrf1 dictates its post-translational modification and its ability to transactivate target genes. Here, we have elucidated that topovectorial mechanisms control the juxtamembrane processing of Nrf1 on the cyto/nucleoplasmic side of endoplasmic reticulum (ER), whereupon it is cleaved and degraded to remove various lengths of its N-terminal domain (NTD, also refold into a UBL module) and acidic domain-1 (AD1) to yield multiple isoforms. Notably, an N-terminal ∼12.5-kDa polypeptide of Nrf1 arises from selective cleavage at an NHB2-adjoining region within NTD, whilst other longer UBL-containing isoforms may arise from proteolytic processing of the protein within AD1 around PEST1 and Neh2L degrons. The susceptibility of Nrf1 to proteolysis is determined by dynamic repositioning of potential UBL-adjacent degrons and cleavage sites from the ER lumen through p97-driven retrotranslocation and -independent pathways into the cyto/nucleoplasm. These repositioned degrons and cleavage sites within NTD and AD1 of Nrf1 are coming into their bona fide functionality, thereby enabling it to be selectively processed by cytosolic DDI-1/2 proteases and also degraded via 26S proteasomes. The resultant proteolytic processing of Nrf1 gives rise to a mature ∼85-kDa CNC-bZIP transcription factor, which regulates transcriptional expression of cognate target genes. Furthermore, putative ubiquitination of Nrf1 is not a prerequisite necessary for involvement of p97 in the client processing. Overall, the regulated juxtamembrane proteolysis (RJP) of Nrf1, though occurring in close proximity to the ER, is distinctive from the mechanism that regulates the intramembrane proteolytic (RIP) processing of ATF6 and SREBP1.

A naturally-occurring dominant-negative competitor of Keap1 against its inhibition of Nrf2

ABSTRACTTranscription factor Nrf2 is a master regulator of antioxidant and/or electrophile response elements (AREs/EpREs)driven genes involved in homeostasis, detoxification and adaptation to various stresses. The cytoprotective activity of Nrf2, though being oppositely involved in both cancer prevention and progression, is critically controlled by Keap1 (Kelch-like ECH-associated protein 1) as an adaptor subunit of Cullin 3-based E3 ubiquitin ligase, that is a key sensor for oxidative and electrophilic stresses. Now, we first report a novel naturally-occurring mutant of Keap1, designated Keap1ΔC, which lacks most of its C-terminal Nrf2-interacting domain essential for inhibition of the CNC-bZIP factor. This mutant Keap1ΔC is yielded by translation from an alternatively mRNA-spliced variant lacking the fourth and fifth exons, but their coding sequences are retained in the wild-type Keap1 locus (with no genomic deletions). Although this variant was found primarily in the human highly-metastatic hepatoma (MHCC97H) cells, it was widely expressed at very lower levels in all other cell lines examined. No matter whether Keap1ΔC retains less or no ability to inhibit Nrf2, it functions as a dominant-negative competitor of Keap1 against its inhibition of Nrf2-target genes. This is due to its antagonist effect on Keap1-mediated turnover of Nrf2 protein.