Biology subjects

Qiu, L.

Publications and source records attributed to Qiu, L..

99 records · Page 6Linked to original sources

bioRxiv2018

Coordinative Metabolism of Glutamine Carbon and Nitrogen in proliferating Cancer Cells Under Hypoxia

Under hypoxia, most of glucose is converted to secretory lactate, which leads to the lack of carbon source from glucose and thus the overuse of glutamine-carbon. However, under such a condition how glutamine nitrogen is disposed to avoid releasing potentially toxic ammonia remains to be determined. Here we identify a metabolic flux of glutamine to secretory dihydroorotate under hypoxia. We found that glutamine nitrogen is indispensable to nucleotide biosynthesis, but enriched in dihyroorotate and orotate rather than processing to its downstream uridine monophosphate under hypoxia. Dihyroorotate, not orotate, is then secreted out of cells. The specific metabolic pathway occurs in vivo and is required for tumor growth. Such a metabolic pathway renders glutamine mainly to acetyl coenzyme A for lipogenesis, with the rest carbon and nitrogen being safely removed. Our results reveal how glutamine carbon and nitrogen are coordinatively metabolized under hypoxia, and provide a comprehensive understanding on glutamine metabolism.\n\nSignificanceTumor cells often addict to glutamine, and particularly utilize its carbon for lipogenesis under hypoxia. We reveal that tumor cells package the excessive glutamine-nitrogen into secretory dihydroorotate, instead of toxic ammonia. This specifically reprogrammed pathway supports in vivo tumor growth, and could offer diagnostic markers and therapeutic targets for cancers.

biochemistry↗

bioRxiv2018

Distinct isoforms of Nrf1 diversely regulate different subsets of its cognate target genes

The single Nrf1 gene has capability to be differentially transcripted alongside with alternative mRNA-splicing and subsequent translation through different initiation signals so as to yield distinct lengths of polypeptide isoforms. Amongst them, three of the most representatives are Nrf1, Nrf1{beta} and Nrf1{gamma}, but the putative specific contribution of each isoform to regulating ARE-driven target genes remains unknown. To address this, we have here established three cell lines on the base of the Flp-In T-REx system, which are allowed for tetracycline-inducibly stable expression of Nrf1, Nrf1{beta} and Nrf1{gamma}. The RNA-Sequencing results have demonstrated that a vast majority of differentially expressed genes (i.e. >90% DEGs detected) were dominantly up-regulated by Nrf1 and/or Nrf1{beta} following induction by tetracycline. By contrast, other DEGs regulated by Nrf1{gamma} were far less than those regulated by Nrf1/{beta} (i.e. ~11% of Nrf1 and 7% of Nrf1{beta}). Further transcriptomic analysis revealed that tetracycline-induced expression of Nrf1{gamma} significantly increased the percentage of down-regulated genes in total DEGs. These statistical data were further validated by quantitative real-time PCR. The experimental results indicate that distinct Nrf1 isoforms make diverse and even opposing contributions to regulating different subsets of target genes, such as those encoding 26S proteasomal subunits and others involved in various biological processes and functions. Collectively, Nrf1{gamma} acts as a major dominant-negative competitor against Nrf1/{beta} activity, such that a number of DEGs regulated by Nrf1/{beta} are counteracted by Nrf1{gamma}.

molecular biology↗

bioRxiv2018

Nrf1D is the first candidate secretory transcription factor in the blood plasma, with its precursor existing as a unique redox-sensitive transmembrane CNC-bZIP protein in somatic tissues

Amongst multiple distinct isoforms, Nrf1D is synthesized from translation of an alternatively-spliced transcript of Nrf1 mRNA, with a naturally-occurring deletion of its stop codon-flanking 1466 nucleotides. This molecular event leads to the reading frameshift mutation, which results in a constitutive substitution of the intact Nrf1s C-terminal 72 amino acids (aa, covering the second half of the leucine zipper motif to C-terminal Neh3L domain) by an additional extended 80-aa stretch to generate a unique variant Nrf1D. The C-terminal extra 80-aa region of Nrf1D was identified to fold into a redox-sensitive transmembrane domain that enables it to be tightly integrated within the endoplasmic reticulum (ER) membranes. Notably, the salient feature of Nrf1D confers it to be distinguishable from prototypic Nrf1, such that Nrf1D is endowed with only a less ability than wild-type Nrf1 at mediating target gene expression. Further evidence has been presented revealing that both mRNA and protein levels of Nrf1D were detected to varying extents in somatic tissues. Surprisingly, we also found the existence of Nrf1D-derived isoforms in the blood plasma, implying that it is a candidate secretory transcription factor, although its precursor acts as an integral transmembrane-bound CNC-bZIP protein that entails dynamic topologies, before being unleashed from the ER to enter the blood plasma.

cell biology↗

bioRxiv2018

The Doublesex sex determination pathway regulates reproductive division of labor in honey bees

Eusociality, the ultimate level of social organization, requires reproductive division of labor, and a sophisticated system of communication to maintain societal homeostasis. Reproductive division of labor is maintained by physiological differences between reproductive and sterile castes, typically dictated by pheromonal queen fertility signals that suppress worker reproduction. Intriguingly, reproduction and pheromonal signalling share regulatory machinery across insects.The gene Doublesex (Dsx) controls somatic sex determination and differentiation, including the development of ovaries and secondary sexual characteristics, such as pheromonal signalling. We hypothesized that this regulatory network was co-opted during eusocial evolution to regulate reproductive division of labor. Taking advantage of the breakdown in reproductive division of labor that occurs in honey bees when workers commence to lay eggs in the absence of a queen, we knocked down Dsx to observe effects on ovary development and fertility signal production. As expected, treated workers had lower levels of egg yolk protein, for which Dsx is a cis-regulatory enhancer in other insects, and greatly reduced ovary development. Also as expected, while control workers increased their levels of pheromonal fertility signals, treated workers did not, confirming the role of Dsx in regulating pheromone biosynthesis. We further found that Dsx is part of a large network enriched for regulatory proteins, which is also involved during early larval development, and upregulated in queen-destined larvae. Thus, the ancient developmental framework controlling sex specification and reproduction in solitary insects has been exapted for eusociality, forming the basis for reproductive division of labor and pheromonal signalling pathways.\n\nSignificance statementComplex social insect societies rely on division of reproductive labor among their members. Reproductive individuals ( queens) suppress worker reproduction using pheromonal fertility signalling. We show that an ancient regulatory network that controls specification of sex and secondary sexual characteristics in solitary insects, has been co-opted for both both pheromonal signalling and ovary inactivation in honey bees. In addition, this network is also active during caste specification that takes place during the first few days of larval life. These results show that pheromonal signalling and ovary development share a common regulatory framework, potentially explaining why fertility signalling is honest. Furthermore, they show that higher levels of biological complexity can arise by rewiring and elaborating ancestral gene regulatory networks.

genetics↗

bioRxiv2018

Molecular mechanisms controlling the multistage post-translational processing of endogenous Nrf1α/TCF11 proteins to yield distinct proteoforms within the coupled positive and negative feedback circuits

In an attempt to terminate the chaotic state of the literature on Nrf1/TCF11 with various confused molecular masses, we herein establish a generally acceptable criterion required for identification of its endogenous full-length proteins and derivative isoforms expressed differentially in distinct experimental cell lines. Further work has been focused on the molecular mechanisms that dictate the successive multistate post-translational modifications (i.e. glycosylation by OST, deglycosylation by NGLY, and ubiquitination by Hrd1) of this CNC-bZIP protein and its proteolytic processing to yield multiple isoforms. Several lines of experimental evidence have demonstrated that the nascent Nrf1/TCF11 polypeptide (non-glycosylated) is transiently translocated into the endoplasmic reticulum (ER), in which it becomes an inactive glycoprotein-A, and also folded in a proper topology within and around membranes. Thereafter, dynamic repositioning of the ER-resident domains in Nrf1 glycoprotein is driven by p97-fueled retrotranslocation into extra-ER compartments. Therein, glycoprotein of Nrf1 is allowed for digestion into a deglycoprotein-B and then its progressive proteolytic processing by cytosolic DDI-1/2 and proteasomes to yield distinct proteoforms (i.e. protein-C/D). The processing is accompanied by removal of a major N-terminal ~12.5-kDa polypeptide from Nrf1. Interestingly, our present study has further unraveled that coupled positive and negative feedback circuits exist between Nrf1 and its cognate target genes, including those encoding its regulators p97, Hrd1, DDI-1 and proteasomes. These key players are differentially or even oppositely involved in diverse cellular signalling responses to distinct extents of ER-derived proteotoxic and oxidative stresses induced by different concentrations of proteasomal inhibitors.

biochemistry↗

bioRxiv2018

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

The 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.

biochemistry↗

bioRxiv2018

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

Transcription 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 Keap1{Delta}C, which lacks most of its C-terminal Nrf2-interacting domain essential for inhibition of the CNC-bZIP factor. This mutant Keap1{Delta}C 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 Keap1{Delta}C 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.

molecular biology↗

bioRxiv2018

Nach is a novel ancestral subfamily of the CNC-bZIP transcription factors selected during evolution from the marine bacteria to human

All living organisms have undergone the evolutionary selection under the changing natural environments to survive as diverse life forms. All life processes including normal homeostatic development and growth into organismic bodies with distinct cellular identifications, as well as their adaptive responses to various intracellular and environmental stresses, are tightly controlled by signaling of transcriptional networks towards regulation of cognate genes by many different transcription factors. Amongst them, one of the most conserved is the basic-region leucine zipper (bZIP) family. They play vital roles essential for cell proliferation, differentiation and maintenance in complex multicellular organisms. Notably, an unresolved divergence on the evolution of bZIP proteins is addressed here. By a combination of bioinformatics with genomics and molecular biology, we have demonstrated that two of the most ancestral family members classified into BATF and Jun subgroups are originated from viruses, albeit expansion and diversification of the bZIP superfamily occur in different vertebrates. Interestingly, a specific ancestral subfamily of bZIP proteins is identified and also designated Nach (Nrf and CNC homology) on account of their highly conservativity with NF-E2 p45 subunit-related factors Nrf1/2. Further experimental evidence reveals that Nach1/2 from the marine bacteria exerts distinctive functions from Nrf1/2 in the transcriptional ability to regulate antioxidant response element (ARE)-driven cytoprotective genes. Collectively, an insight into Nach/CNC-bZIP proteins provides a better understanding of distinct biological functions between these factors selected during evolution from the marine bacteria to human.\n\nSignificanceWe identified the novel ancestral subfamily (i.e. Nach) of CNC-bZIP transcription factors with highly conservativity from marine bacteria to human. Combination of bioinformatics with genomics and molecular biology demonstrated that two of the most ancestral family members classified into BATF and Jun subgroups are originated from viruses. The Jun and CNC subfamilies also share a common origin of these bZIP proteins. Further experimental evidence reveals that Nach1/2 from the marine bacteria exerts nuance functions from human Nrf1/2 in the transcriptional ability to regulate antioxidant response element (ARE)-driven genes, responsible for the host cytoprotection against inflammation and cancer. Overall, this study is of multidisciplinary interests to provide a better understanding of distinct biological functions between Nach/CNC-bZIPs selected during evolution.

ecology↗

bioRxiv2017

The Neural System of Metacognition Accompanying Decision-Making in The Prefrontal Cortex

Decision-making is usually accompanied by metacognition, through which a decision maker monitors the decision uncertainty and consequently revises the decision, even prior to feedback. However, the neural mechanisms of metacognition remain controversial: one theory proposes that metacognition coincides the decision-making process; and another addresses that it entails an independent neural system in the prefrontal cortex (PFC). Here we devised a novel paradigm of \"decision-redecision\" to investigate the metacognition process in redecision, in comparison with the decision process. We here found that the anterior PFC, including dorsal anterior cingulate cortex (dACC) and lateral frontopolar cortex (lFPC), were exclusively activated after the initial decisions. dACC was involved in decision uncertainty monitoring, whereas lFPC was involved in decision adjustment controlling, subject to control demands of the tasks. Our findings support that the PFC is essentially involved in metacognition and further suggest that functions of the PFC in metacognition are dissociable.

neuroscience↗