Differential regulation of full-length genome and a single-stranded 7S DNA along the cell cycle in human mitochondria.

Mammalian mitochondria contain full-length genome and a single-stranded 7S DNA. Although the copy number of mitochondrial DNA (mtDNA) varies depending on the cell type and also in response to diverse environmental stresses, our understanding of how mtDNA and 7S DNA are maintained and regulated is limited, partly due to lack of reliable in vitro assay systems that reflect the in vivo functionality of mitochondria. Here we report an in vitro assay system to measure synthesis of both mtDNA and 7S DNA under a controllable in vitro condition.

Regulation of inter- and intramolecular interaction of RNA, DNA, and proteins by MLE.

Drosophila maleless (MLE) is a member of helicase superfamily 2 and functions as a dosage compensation factor essential for the development of male flies. This function provides a good opportunity to investigate diverse biochemical activities associated with MLE in the context of a defined in vivo pathway, i.e.

Regulation of the catalytic function of topoisomerase II alpha through association with RNA.

Topoisomerase IIalpha interacts with numerous nuclear factors, through which it is engaged in diverse nuclear events such as DNA replication, transcription and the formation or maintenance of heterochromatin. We previously reported that topoisomerase IIalpha interacts with RNA helicase A (RHA), consistent with a recent view that topoisomerases and helicases function together. Intrigued by our observation that the RHA-topoisomerase IIalpha interaction is sensitive to ribonuclease A, we explored whether the RHA-topoisomerase IIalpha interaction can be recapitulated in vitro using purified proteins and a synthetic RNA.

Nuclear factor 45 (NF45) is a regulatory subunit of complexes with NF90/110 involved in mitotic control.

Nuclear factor 90 (NF90) and its C-terminally extended isoform, NF110, have been isolated as DNA- and RNA-binding proteins together with the less-studied protein NF45. These complexes have been implicated in gene regulation, but little is known about their cellular roles and whether they are redundant or functionally distinct. We show that heterodimeric core complexes, NF90-NF45 and NF110-NF45, exist within larger complexes that are more labile and contain multiple NF90/110 isoforms and additional proteins.

Transcriptional regulation of multi-drug tolerance and antibiotic-induced responses by the histone-like protein Lsr2 in M. tuberculosis.

Multi-drug tolerance is a key phenotypic property that complicates the sterilization of mammals infected with Mycobacterium tuberculosis. Previous studies have established that iniBAC, an operon that confers multi-drug tolerance to M. bovis BCG through an associated pump-like activity, is induced by the antibiotics isoniazid (INH) and ethambutol (EMB).

MLE functions as a transcriptional regulator of the roX2 gene.

Dosage compensation is a process that equalizes transcription activity between the sexes. In Drosophila, two non-coding RNA, roX1 and roX2, and at least six protein regulators, MSL-1, MSL-2, MSL-3, MLE, MOF, and JIL-1, have been identified as essential for dosage compensation. Although there is accumulating evidence of the intricate functional and physical interactions between protein and RNA regulators, little is known about how roX RNA expression and function are modulated in coordination with protein regulators.

A functional interaction between RHA and Ubc9, an E2-like enzyme specific for Sumo-1.

RNA helicase A (RHA) is a member of the DEAH helicase family of proteins. Recent studies imply the role of RHA in the regulation of the topology of chromatin DNA, which could influence diverse nuclear processes such as transcription activity of the chromatin DNA and chromosome condensation. We previously reported that Ubc9, an E2-like enzyme specific for small ubiquitin-like modifier 1 (Sumo-1), is required for the interaction between RHA and topoisomerase IIalpha.

Identification of toposome, a novel multisubunit complex containing topoisomerase IIalpha.

Topoisomerase IIalpha plays essential roles in chromosome segregation. However, it is not well understood how topoisomerase IIalpha exerts its function during mitosis. In this report, we find that topoisomerase IIalpha forms a multisubunit complex, named toposome, containing two ATPase/helicase proteins (RNA helicase A and RHII/Gu), one serine/threonine protein kinase (SRPK1), one HMG protein (SSRP1), and two pre-mRNA splicing factors (PRP8 and hnRNP C).

Selective regulation of gene expression by nuclear factor 110, a member of the NF90 family of double-stranded RNA-binding proteins.

Members of the nuclear factor 90 (NF90) family of double-stranded RNA (dsRNA)-binding proteins have been implicated in several biological processes including the regulation of gene expression. cDNA sequences predict that the proteins have a functional nuclear localization signal and two dsRNA-binding motifs (dsRBMs), and are identical at their N termini. Isoforms are predicted to diverge at their C termini as well as by the insertion of four amino acid residues (NVKQ) between the two dsRBMs.

DEN1 is a dual function protease capable of processing the C terminus of Nedd8 and deconjugating hyper-neddylated CUL1.

Nedd8 activates ubiquitination by increasing the efficiency of polyubiquitin chain assembly through its covalent conjugation to cullin molecules. Here we report the isolation, cloning, and characterization of a novel human Nedd8-specific protease called DEN1. Human DEN1 is encoded by AAH31411.

RNA helicase A interacts with dsDNA and topoisomerase IIalpha.

RNA helicase A (RHA) is a multifunctional protein involved in various nuclear processes such as transcription and RNA export. It is believed that the interacting factors play important roles in determining the functional specificity of RHA. Here we show that RHA directly interacts with double-stranded (ds) nucleic acids (NAs) and assembles complexes with topoisomerase IIalpha.

The growth factor granulin interacts with cyclin T1 and modulates P-TEFb-dependent transcription.

Cyclin T1, together with the kinase CDK9, is a component of the transcription elongation factor P-TEFb which binds the human immunodeficiency virus type 1 (HIV-1) transactivator Tat. P-TEFb facilitates transcription by phosphorylating the carboxy-terminal domain (CTD) of RNA polymerase II. Cyclin T1 is an exceptionally large cyclin and is therefore a candidate for interactions with regulatory proteins.

Effect of an exogenous trigger on the pathogenesis of lupus in (NZB x NZW)F1 mice.

Objective: This study examined the interactions between exogenous and endogenous factors shaping the phenotype of lupus in autoimmune (NZB x NZW)F(1) mice exposed to pristane, a model environmental trigger.

Methods: Frequencies of various autoantibodies in untreated NZB/NZW mice were determined by various means (immunoprecipitation, enzyme-linked immunosorbent assay [ELISA], Crithidia luciliae kinetoplast staining). Pristane or saline was administered intraperitoneally to 9-12-week-old NZB/NZW mice, followed by serial studies of autoantibodies, total Ig levels (ELISA), and proteinuria (dipstick).

RH70, a bidirectional RNA helicase, co-purifies with U1snRNP.

In this report, I describe the co-purification of a novel 70-kDa RNA helicase (RH70) and U1snRNP through six column steps. Peptide sequence analysis by mass spectrometry and Edman degradation revealed that RH70 is the previously reported DDX17. Biochemical characterization of RH70, obtained by partial separation from U1snRNP, yielded the following results.

Gene expression profile of HIV-1 Tat expressing cells: a close interplay between proliferative and differentiation signals.

Background: Expression profiling holds great promise for rapid host genome functional analysis. It is plausible that host expression profiling in an infection could serve as a universal phenotype in virally infected cells. Here, we describe the effect of one of the most critical viral activators, Tat, in HIV-1 infected and Tat expressing cells.