Niall S. Kenneth

Regulation of hypoxia-inducible factor-1α by NF-κB

HIF (hypoxia-inducible factor) is the main transcription factor activated by low oxygen tensions. HIF-1α (and other α subunits) is tightly controlled mostly at the protein level, through the concerted action of a class of enzymes called PHDs (prolyl hydroxylases) 1, 2 and 3. Most of the knowledge of HIF derives from studies following hypoxic stress; however, HIF-1α stabilization is also found in non-hypoxic conditions through an unknown mechanism. In the present study, we demonstrate that NF-κB (nuclear factor κB) is a direct modulator of HIF-1α expression. The HIF-1α promoter is responsive to selective NF-κB subunits. siRNA (small interfering RNA) studies for individual NF-κB members revealed differential effects on HIF-1α mRNA levels, indicating that NF-κB can regulate basal HIF-1α expression. Finally, when endogenous NF-κB is induced by TNFα (tumour necrosis factor α) treatment, HIF-1α levels also change in an NF-κB-dependent manner. In conclusion, we find that NF-κB can regulate basal TNFα and, in certain circumstances, the hypoxia-induced HIF-1α.

Regulation of hypoxia inducible factor-1? by NF-?B.

HIF (hypoxia-inducible factor) is the main transcription factor activated by low oxygen tensions. HIF-1α (and other α subunits) is tightly controlled mostly at the protein level, through the concerted action of a class of enzymes called PHDs (prolyl hydroxylases) 1, 2 and 3. Most of the knowledge of HIF derives from studies following hypoxic stress; however, HIF-1α stabilization is also found in non-hypoxic conditions through an unknown mechanism. In the present study, we demonstrate that NF-κB (nuclear factor κB) is a direct modulator of HIF-1α expression. The HIF-1α promoter is responsive to selective NF-κB subunits. siRNA (small interfering RNA) studies for individual NF-κB members revealed differential effects on HIF-1α mRNA levels, indicating that NF-κB can regulate basal HIF-1α expression. Finally, when endogenous NF-κB is induced by TNFα (tumour necrosis factor α) treatment, HIF-1α levels also change in an NF-κB-dependent manner. In conclusion, we find that NF-κB can regulate basal TNFα and, in certain circumstances, the hypoxia-induced HIF-1α.

Regulation of gene expression by hypoxia

Hypoxia induces profound changes in the cellular gene expression profile. The discovery of a major transcription factor family activated by hypoxia, HIF (hypoxia-inducible factor), and the factors that contribute to HIF regulation have greatly enhanced our knowledge of the molecular aspects of the hypoxic response. However, in addition to HIF, other transcription factors and cellular pathways are activated by exposure to reduced oxygen. In the present review, we summarize the current knowledge of how additional hypoxia-responsive transcription factors integrate with HIF and how other cellular pathways such as chromatin remodelling, translation regulation and microRNA induction, contribute to the co-ordinated cellular response observed following hypoxic stress.

RETRACTED: Elevated tRNAiMet Synthesis Can Drive Cell Proliferation and Oncogenic Transformation

Characteristics of transformed and tumor cells include increased levels of protein synthesis and elevated expression of RNA polymerase (pol) III products, such as tRNAs and 5S rRNA. However, whether deregulated pol III transcription contributes to transformation has been unclear. Generating cell lines expressing an inducible pol III-specific transcription factor, Brf1, allowed us to raise tRNA and 5S rRNA levels specifically. Brf1 induction caused an increase in cell proliferation and oncogenic transformation, whereas depletion of Brf1 impeded transformation. Among the gene products induced by Brf1 is the tRNA(iMet) that initiates polypeptide synthesis. Overexpression of tRNA(iMet) is sufficient to stimulate cell proliferation and allow immortalized fibroblasts to form foci in culture and tumors in mice. The data indicate that elevated tRNA synthesis can promote cellular transformation.

Direct Regulation of RNA Polymerase III Transcription by RB, p53 and c-Myc

The synthesis of tRNA and 5S rRNA by RNA polymerase (pol) III is cell cycle regulated in higher organisms. Overexpression of pol III products is a general feature of transformed cells. These observations may be explained by the fact that a pol III-specific transcription factor, TFIIIB, is strongly regulated by the tumour suppressors RB and p53, as well as the proto-oncogene product c-Myc. RB and p53 repress TFIIIB, but this restraint can be lost in tumours through a variety of mechanisms. In contrast, c-Myc binds and activates TFIIIB, causing potent induction of pol III transcription. Using chromatin immunoprecipitation and RNA interference, we show that c-Myc interacts with tRNA and 5S rRNA genes in transformed cervical cells, stimulating their expression. Availability of pol III products may be an important determinant of a cells capacity to grow. The ability to regulate pol III output may therefore be integral to the growth control functions of RB, p53 and c-Myc.

SWI/SNF Regulates the Cellular Response to Hypoxia

Hypoxia induces a variety of cellular responses such as cell cycle arrest, apoptosis, and autophagy. Most of these responses are mediated by the hypoxia-inducible factor-1α. To induce target genes, hypoxia-inducible factor-1α requires a chromatin environment conducive to allow binding to specific sequences. Here, we have studied the role of the chromatin-remodeling complex SWI/SNF in the cellular response to hypoxia. We find that SWI/SNF is required for several of the cellular responses induced by hypoxia. Surprisingly, hypoxia-inducible factor-1α is a direct target of the SWI/SNF chromatin-remodeling complex. SWI/SNF components are found associated with the hypoxia-inducible factor-1α promoter and modulation of SWI/SNF levels results in pronounced changes in hypoxia-inducible factor-1α expression and its ability to transactivate target genes. Furthermore, impairment of SWI/SNF function renders cells resistant to hypoxia-induced cell cycle arrest. These results reveal a previously uncharacterized dependence of hypoxia signaling on the SWI/SNF complex and demonstrate a new level of control over the hypoxia-inducible factor-1α system.

TRRAP and GCN5 are used by c-Myc to activate RNA polymerase III transcription

Activation of RNA polymerase (pol) II transcription by c-Myc generally involves recruitment of histone acetyltransferases and acetylation of histones H3 and H4. Here, we describe the mechanism used by c-Myc to activate pol III transcription of tRNA and 5S rRNA genes. Within 2 h of its induction, c-Myc appears at these genes along with the histone acetyltransferase GCN5 and the cofactor TRRAP. At the same time, occupancy of the pol III-specific factor TFIIIB increases and histone H3 becomes hyperacetylated, but increased histone H4 acetylation is not detected at these genes. The rapid acetylation of histone H3 and promoter assembly of TFIIIB, c-Myc, GCN5, and TRRAP are followed by recruitment of pol III and transcriptional induction. The selective acetylation of histone H3 distinguishes pol III activation by c-Myc from mechanisms observed in other systems.

Evolutionary Conserved Regulation of HIF-1β by NF-κB

Hypoxia Inducible Factor-1 (HIF-1) is essential for mammalian development and is the principal transcription factor activated by low oxygen tensions. HIF-α subunit quantities and their associated activity are regulated in a post-translational manner, through the concerted action of a class of enzymes called Prolyl Hydroxylases (PHDs) and Factor Inhibiting HIF (FIH) respectively. However, alternative modes of HIF-α regulation such as translation or transcription are under-investigated, and their importance has not been firmly established. Here, we demonstrate that NF-κB regulates the HIF pathway in a significant and evolutionary conserved manner. We demonstrate that NF-κB directly regulates HIF-1β mRNA and protein. In addition, we found that NF-κB–mediated changes in HIF-1β result in modulation of HIF-2α protein. HIF-1β overexpression can rescue HIF-2α protein levels following NF-κB depletion. Significantly, NF-κB regulates HIF-1β (tango) and HIF-α (sima) levels and activity (Hph/fatiga, ImpL3/ldha) in Drosophila, both in normoxia and hypoxia, indicating an evolutionary conserved mode of regulation. These results reveal a novel mechanism of HIF regulation, with impact in the development of novel therapeutic strategies for HIF–related pathologies including ageing, ischemia, and cancer.

An inactivating caspase 11 passenger mutation originating from the 129 murine strain in mice targeted for c-IAP1

A recent study revealed that ES (embryonic stem) cell lines derived from the 129 murine strain carry an inactivating mutation within the caspase 11 gene (Casp4) locus [Kayagaki, Warming, Lamkanfi, Vande Walle, Louie, Dong, Newton, Qu, Liu, Heldens, Zhang, Lee, Roose-Girma and Dixit (2011) Nature 479, 117–121]. Thus, if 129 ES cells are used to target genes closely linked to caspase 11, the resulting mice might also carry the caspase 11 deficiency as a passenger mutation. In the present study, we examined the genetic loci of mice targeted for the closely linked c-IAP (cellular inhibitor of apoptosis) genes, which were generated in 129 ES cells, and found that, despite extensive backcrossing into a C57BL/6 background, c-IAP1−/− animals are also deficient in caspase 11. Consequently, data obtained from these mice should be re-evaluated in this new context.

IAP proteins: regulators of cell migration and development.

The cytoprotective properties of vertebrate inhibitor of apoptosis (IAP) proteins have been the subject of much study. These proteins have, however, emerged as key signaling intermediates modulating a variety of cellular functions through their ability to act as E3 ubiquitin ligases. This review will focus on the cell death-independent roles of the IAP proteins, focusing on recent reports indicating that c-IAPs and XIAP are key molecules involved in modulating cell migration and development.