Yan Lavrovsky

Role of redox-regulated transcription factors in inflammation, aging and age-related diseases

A progressive rise of oxidative stress due to the altered redox homeostasis appears to be one of the hallmarks of the aging process. Reactive oxygen species (ROS) also serve as signaling agents for inflammation, a systemic defensive reaction against microbial pathogens and other foreign bodies. Changes in the pattern of gene expression through ROS-sensitive transcription factors give rise to both aging and inflammation phenotypes. Chronic oxidative stress and inflammatory reaction also lead to many age-associated diseases such as atherosclerosis and arthritis. Transcription factors that are directly influenced by ROS and proinflammatory cytokines include nuclear factor kappa B (NF-kappaB), activator protein 1 (AP-1), specificity protein 1 (Sp1), peroxisome proliferator-activated receptors (PPARs) and other members of the nuclear receptor superfamily. Here we describe the basic components of the intracellular redox control machinery and their dysregulation with age leading to altered transcription factor function and age-associated pathophysiology.

Regulation of Androgen Action

Publisher Summary This chapter describes the different aspects of regulation of androgen action. Androgens belong to a class of C-19 steroids secreted primarily by the testis and adrenal cortex. Hormonally active androgens promote reproductive and anabolic functions. Both reproductive and anabolic effects of androgens are mediated by their interaction with the androgen receptor (AR), a member of the steroid-thyroid hormone-retinoid-vitamin D superfamily of nuclear receptors (NRs) that function as ligand-activated transcription factors. Almost all of the androgen functions, except its conversion to estrogen by the enzyme aromatase in certain target cells are known to be mediated by the androgen receptor. The androgen receptor are coded by a single copy gene, which is located on the X chromosome. The functional relevance of the segmented domain structure of the NR superfamily is supported by the results of deletion mutagenesis and domain swapping among various receptor proteins. The interaction between the amino terminal and the steroid-binding carboxy-terminal end in the AR transactivation function was initially indicated by the finding that a segment within the hormone-binding domain exerts an inhibitory influence in the transcription regulatory activity of the AR, and the deletion of this region results in ligand-independent activation of the receptor.

Androgen Receptor: Structural Domains and Functional Dynamics after Ligand-Receptor Interaction

Abstract: Androgens are C‐19 steroids secreted primarily from the testes and adrenals that play a critical role in reproduction. Reproductive functions of androgens are mediated through coordination of diverse physiological processes ranging from brain functions to specific cell proliferation and apoptosis. At the molecular level, most of these regulatory influences are exerted by altered expression of appropriate genes by the androgen receptor (AR), a member of the nuclear receptor (NR) superfamily. The unliganded AR is a cytoplasmic protein and, upon ligand binding, it translocates into the nucleus. Thereafter, in conjunction with other transcription factors and coactivators, the AR influences transcription of target genes through a multistep process that includes its clustering in a subnuclear compartment. Here, we describe the genomic organization of the AR, the role of individual structural domains in specific AR function, and the influence of agonistic/antagonistic ligands in the intracellular movement of the receptor. We also show that the AR is capable of undergoing multiple rounds of nucleocytoplasmic recycling after ligand binding and dissociation. Xenobiotic ligands, considered as selective androgen receptor modulators (SARMs), can modulate AR activity by inhibiting either its nuclear translocation or its subnuclear clustering and subsequent transactivation function.

Age-dependent increase of heme oxygenase–1 gene expression in the liver mediated by NFκB

Heme, the iron-porphyrin coordination complex, released from the degradation of hemoproteins, is a strong prooxidant. It is enzymatically degraded by heme oxygenase to free iron, carbon monoxide and biliverdin. Biliverdin and its reduced metabolite bilirubin are two potent physiological antioxidants. Here we show a progressive increase of steady-state levels of the mRNA encoding the inducible isoform of this enzyme (heme oxygenase-1) in the rat liver during aging. We had previously reported that aging is associated with increased activation of the nuclear factor kappaB (NFkappaB). We now provide evidence to establish that overexpression of NFkappaB in transfected liver-derived HepG2 cells can cause a marked induction of the endogenous heme oxygenase-1 (HO-1) mRNA and activation of the cotransfected HO-1 gene promoter. Taken together, these results support the conclusion that enhanced oxidative stress during aging is accompanied by compensatory induction of the antioxidant enzyme HO-1 through activation of the NFkappaB pathway.

Heme oxygenase-1 in tissue pathology: The Yin and Yang

Heme is a versatile molecule in nature and serves as the prosthetic moiety for numerous hemoproteins involved in oxygen delivery, electron transfer, and signal transduction. 1 However, when left unattended, the same heme can promote free radical formation and lipid peroxidation, resulting in cell damage and tissue injury. 2-4 Thus, a fine balance between heme synthesis and catabolism is essential for the maintenance of cellular homeostasis. Whereas biosynthesis of heme is catalyzed by multiple enzymes, the only physiological mechanism of heme degradation is through heme oxygenase (HO). 1 Heme oxygenase catalyzes breakdown of the protoporphyrin ring, producing biliverdin, carbon monoxide, and free ferrous iron. 5 To this date, three isoforms of heme oxygenase have been identified. 6 Among them, HO-1, unlike the other two (HO-2 and -3), shows limited expression under normal situations and is induced by a variety of physiological stimuli. 5,7,8 Heme, free iron, and a number of oxidative stressors can markedly potentiate this inductive response. Transcriptional control of HO-1 is mediated by multiple factors including NFκB and AP-1. Both of these transcription factors are activated by free radicals generated by heme, iron, and other unrelated agents. 5,7-10 Then what is the biological meaning of the specific induction? For some time, evidence has been accumulating to suggest that HO-1 induction is an adaptive response to cellular stresses. 5,7,8

A rapid and reliable PCR-based assay for gene transmission and sex determination in newborn transgenic mice.

This article describes a reliable and rapid method for simultaneous detection of a transgene and sex determination in the newborn mouse pups by PCR using three sets of primers in a single reaction. One set of sense/antisense primers is used to amplify the experimental transgene (androgen receptor gene in this case), the second set for the mouse Y-chromosome-specific SRY gene, and the third set for the β subunit of the thyroid stimulating hormone (TSHβ), an internal control. This procedure allowed us to promptly analyze pups born from transgenic founders carrying the androgen receptor transgene and, at the same time, establish the sex of the animals. The method is simple, rapid and highly reproducible.

Ribozyme Technology and Drug Development

Publisher Summary This chapter reviews the state of knowledge of the oligonucleotide-based therapeutic approaches with an emphasis on ribozyme technology. Many ribozymes are used for treating viral diseases and neoplastic, cardiovascular, and genetic disorders. Effective use of the antisense oligonucleotides and ribozymes for therapeutic purposes depends on optimization of the targeted delivery, pharmacokinetics, and drug metabolism as well as protection against adverse cytokinetic response. However, rapid advances in this field promise better use of these therapeutic drugs. The chapter discusses nonenzymatic antisense oligonucleotides as inhibitors of specific gene expression. These include peptide nucleic acids (PNAs) and triplex-forming oligonucleotides. RNA based enzymes are described focusing on RNA folding, ribozyme structure and function, and ribozymes of different types and their effects on gene expression regulation in vitro. Catalytic DNAs, therapeutic applications of catalytic oligonucleotides, ribozyme delivery, pharmacokinetics, and metabolism and future prospects are discussed. Therapeutic applications look at application as antiviral agents, treatment of neoplastic diseases, cardiovascular diseases, and genetic disorders.