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PubMed2006

Limited role for the DsrA and RprA regulatory RNAs in rpoS regulation in Salmonella enterica.

RpoS, the sigma factor of enteric bacteria that responds to stress and stationary phase, is subject to complex regulation acting at multiple levels, including transcription, translation, and proteolysis. Increased translation of rpoS mRNA during growth at low temperature, after osmotic challenge, or with a constitutively activated Rcs phosphorelay depends on two trans-acting small regulatory RNAs (sRNAs) in Escherichia coli. The DsrA and RprA sRNAs are both highly conserved in Salmonella enterica, as is their target, an inhibitory antisense element within the rpoS untranslated leader. Analysis of dsrA and rprA deletion mutants indicates that while the increased translation of RpoS in response to osmotic challenge is conserved in S. enterica, dependence on these two sRNA regulators is much reduced. Furthermore, low-temperature growth or constitutive RcsC activation had only modest effects on RpoS expression, and these increases were, respectively, independent of dsrA or rprA function. This lack of conservation of sRNA function suggests surprising flexibility in RpoS regulation.

Bacterial Proteins↗
PubMed2006

Noncoding RNAs in the mammalian central nervous system.

The central nervous system (CNS) is arguably one of the most complex systems in the universe. To understand the CNS, scientists have investigated a variety of molecules, including proteins, lipids, and various small molecules. However, one large class of molecules, noncoding RNAs (ncRNAs), has been relatively unexplored. ncRNAs function directly as structural, catalytic, or regulatory molecules rather than serving as templates for protein synthesis. The increasing variety of ncRNAs being identified in the CNS suggests a strong connection between the biogenesis, dynamics of action, and combinatorial regulatory potential of ncRNAs and the complexity of the CNS. In this review, we give an overview of the diversity and abundance of ncRNAs before delving into specific examples that illustrate their importance in the CNS. In particular, we cover recent evidence for the roles of microRNAs, small nucleolar RNAs, retrotransposons, the NRSE small modulatory RNA, and BC1/BC200 in the CNS. Finally, we speculate why ncRNAs are well adapted to improving organism-environment interactions.

Animals↗
PubMed2004

RNA interference by small hairpin RNAs synthesised under control of the human 7S K RNA promoter.

Small interfering RNAs (siRNAs) represent RNA duplexes of 21 nucleotides in length that inhibit gene expression. We have used the human gene-external 7S K RNA promoter for synthesis of short hairpin RNAs (shRNAs) which efficiently target human lamin mRNA via RNA interference (RNAi). Here we demonstrate that orientation of the target sequence within the shRNA construct is important for interference. Furthermore, effective interference also depends on the length and/or structure of the shRNA. Evidence is presented that the human 7S K promoter is more active in vivo than other gene-external promoters, such as the human U6 small nuclear RNA (snRNA) gene promoter.

HeLa Cells↗
PubMed2004

Expression and function of brain specific small RNAs.

Small non-messenger RNAs (snmRNAs) are a heterogeneous group of non-coding RNAs with a variety of regulatory functions including regulation of protein expression and guidance in RNA modifications. They are actively being investigated in Archaebacteria, yeast, invertebrates and mammals. Brain-specific snmRNAs have been identified in mammals and they seem to contribute to neuronal differentiation during development and to brain functions subserving learning and memory. Here we review the current knowledge of the properties, expression and functions of three groups of brain-specific snmRNAs: small nucleolar RNAs, BC1/BC200 RNAs and microRNAs.

Animals↗
PubMed1999

Two internal sequence elements modulate transcription from the external human 7S K RNA gene promoter in vivo.

Constructs of the external promoter of the human 7S K RNA gene in combination with different reporter elements and varying amounts of 7S K internal sequences were analyzed for efficient transcription by RNA polymerase III (pol III) in vitro and in vivo. In vitro, the 7S K promoter alone (-245 to -1) revealed full activity, compared to the entire wild-type gene. In vivo, however, the activity of the gene-external 7S K promoter, albeit clearly functional by itself, was positively modulated by internal sequence elements. Fusion constructs containing increasing amounts of transcribed 7S K sequences revealed that two elements were responsible for this activation. One element is associated with the initiator region (+1 to +8) of this class III gene. The second sequence comprises the 5' half of a cryptic A-box starting at +10 of the 7S K RNA sequence. In the context of a totally unrelated vector sequence, a GGC element alone was sufficient to functionally replace that cryptic A-box. Thus, it appears that in context of the 7S K RNA gene--and possibly the 7S L and U6 RNA genes as well--structurally divergent A-box-like elements function as internal modulators of these pol III promoters.

Genes, Regulator↗
PubMed1989

Autoimmune response to the Ro/SSA particle is directed to the human antigen.

Autoantibodies to defined cellular antigens in systemic lupus erythematosus (SLE) are usually directed to conserved epitopes on ubiquitous macromolecules including histone, Sm + nRNP (URNP particles), DNA, and La(SSB). We report here that the autoimmune response to the Ro(SSA) RNA protein particle is directed to epitopes on the human antigen which are not conserved in evolution. Ro(SSA) from bovine, rat, and mouse Ro(SSA) particles cross-react with human autoantibodies less effectively than does human Ro(SSA), and antigenically active Ro(SSA) is not detectable in chicken thymus extracts with the assays employed. These data suggest a special role for the Ro(SSA) antigen in the initiation and/or perpetuation of the anti-Ro(SSA) response in autoimmune disease.

Absorption↗
PubMed1997

Molecular analysis of snRNAs and scRNAs using autoantibodies from patients with systemic lupus erythematosus.

Immunoprecipitation analysis of total HeLa cells RNA extract by protein A-Sheparose purified autoantibodies and pCp 32P-3' end labeling RNAs revealed that U1, U2, U4 and U5 snRNAs are related with anti-Sm or U1nRNP autoantibodies, while the hY1, hY3, hY4 and hY5 scRNAs were related to anti-SSA/Ro autoantibodies present in sera of patients with Systemic Lupus Erythematosus. The authors detected molecular snRNAs and scRNAs specificities by autoantibodies in 71 sera, the molecular RNA specificity for anti-Sm (U1, U2, U4 and U5 snRNAs) was present in 39%; anti-SSA/Ro sera reacted against scRNAs (hY1, hY3, hY4 and hY5) in 36%, then anti-U1nRNP sera recognized U1 snRNA in 13% of sera and anti-rRNP related with rRNA were recognized in 8%. Twenty-nine SLE sera were RNA negative. A molecular characterization of the autoantibodies in sera from SLE patients may be a useful tool for clinical and laboratory diagnosis of SLE, and the use of autoantibodies as molecular probes allows to continue exploring some basic mechanism of gene expression.

Autoantibodies↗
PubMed1993

Multiple dispersed loci produce small cytoplasmic Alu RNA.

Alu repeats are short interspersed elements (SINEs) of dimeric structure whose transposition sometimes leads to heritable disorders in humans. Human cells contain a poly(A)- small cytoplasmic transcript of -120 nucleotides (nt) homologous to the left Alu monomer. Although its monomeric size indicates that small cytoplasmic Alu (scAlu) RNA is not an intermediary of human Alu transpositions, a less abundant poly(A)-containing Alu transcript of dimeric size and specificity expected of a transposition intermediary is also detectable in HeLa cells (A. G. Matera, U. Hellmann, M. F. Hintz, and C. W. Schmid, Mol. Cell. Biol. 10:5424-5432, 1990). Although its function is unknown, the accumulation of Alu RNA and its ability to interact with a conserved protein suggest a role in cell biology (D.-Y. Chang and R. J. Maraia, J. Biol. Chem. 268:6423-28, 1993). The relationship between the -120- and -300-nt Alu transcripts had not been determined. However, a B1 SINE produces scB1 RNA by posttranscriptional processing, suggesting a similar pathway for scAlu. An Alu SINE which recently transposed into the neurofibromatosis 1 locus was expressed in microinjected frog oocytes. This neurofibromatosis 1 Alu produced a primary transcript followed by the appearance of the scAlu species. 3' processing of a synthetic -300-nt Alu RNA by HeLa nuclear extract in vitro also produced scAlu RNA. Primer extension of scAlu RNA indicates synthesis by RNA polymerase III. HeLa-derived scAlu cDNAs were cloned so as to preserve their 5'-terminal sequences and were found to correspond to polymerase III transcripts of the left monomeric components of three previously identified Alu SINE subfamilies. Rodent x human somatic cell hybrids express Alu RNAs whose size, heterogeneous length, and chromosomal distribution indicate their derivation from SINEs. The coexpression of dimeric and monomeric Alu RNA in several hybrids suggests a precursor-product relationship.

Animals↗
PubMedDate not supplied

[Nuclear and cytoplasmic RNA showing hybridization with repetitive sequences B1 and B2 of the mouse genome].

Heavy nuclear RNA contains high amounts of transcripts from repetitive sequences B1 and B2. Cytoplasmic poly(A+)RNA and in particular, polysomal poly(A)+RNA (mRNA) also contain these sequences though their content is much lower. An abundant 2 kb mRNA of mouse liver was found to contain B2 sequence. A novel class of RNA is described namely as low molecular weight heterogeneous (200-400 nucleotide long) poly(A)+RNA. These RNAs contain B1 and B2 sequences and are located mostly in the cytoplasm but not in polysomes. Tumor cells contain high amounts of small B2+RNAs. The content of B1+ small cytoplasmic RNA is much lower. The hybridization tests confirm the existence of extended homology between B1 sequence and 4.5 S small nuclear RNA which has been earlier deduced from comparison of their base sequences. Homology of B1 and small cytoplasmic 7S RNA was also detected. No hybridization of B2 to small nuclear or small cytoplasmic RNAs could be detected.

Animals↗
PubMed1998

An E-box sequence acts as a transcriptional activator for BC1 RNA expression by RNA polymerase III in the brain.

BC1 RNA is a small cytoplasmic RNA that is transcribed by RNA polymerase III (Pol III) in the rodent nervous system. In addition to essential intragenic promoter elements for Pol III, the BC1 RNA gene has five E-box sequences (CANNTG) in its 5' flanking region. Deletion analysis using an in vitro transcription system revealed that the region containing the E2 site (CAATTG) was necessary for effective transcription of BC1 RNA. A construct with point mutations within the E2 site showed reduced transcriptional activity. Furthermore, DNaseT I protection and gel retardation assays demonstrated that the E2 site was recognized specifically by a brain nuclear protein(s). These results suggest that the upstream E-box sequence and its binding protein may be involved in the regulation by Pol III of preferential BC1 RNA expression in the brain.

Animals↗
PubMed1997

Expression of neural BC1 RNA: induction in murine tumours.

BC1 RNA is a small cytoplasmic RNA polymerase III transcript that is expressed in the rodent nervous system. The RNA is selectively expressed in neurons where it is located in somatodendritic domains. BC1 RNA is not normally detectable in non-neuronal somatic cells; it is however expressed in germ cells and in cultured immortal cell lines of various non-neural origins. We therefore sought to establish whether the neuron-specific regulation of BC1 expression is altered in non-neural tumour cells. Oncogen and chemical carcinogen induced mouse tumours were analysed for the presence of BC1 RNA, using Northern transfer and in situ hybridisation. Here we report that BC1 RNA is selectively expressed in tumour cells, but not in corresponding normal tissues. These results indicate that neural-specific regulation of BC1 expression is lacking in murine tumour cells of non-neural origin.

Animals↗
PubMedDate not supplied

Transcription, processing and nuclear transport of a B1 Alu RNA species complementary to an intron of the murine alpha-fetoprotein gene.

The Alu sequence family comprises the major dispersed repeat sequences of rodent and primate genomes, numbering greater than 300,000 copies in the human haploid genome. The function of these elements is unknown. The sequences can be transcribed by RNA polymerase III and represent a substantial fraction of total heterogeneous nuclear RNA. Alu sequences can be found both in the flanking regions and within the transcription units of several well-characterized genes. Here we show that some members of the mouse B1 Alu sequence family encode a small cytoplasmic RNA. The mouse B1 sequence is congruent to 130 nucleotides long and shows homology with the monomeric units of the dimeric 300-nucleotide primate sequence. By means of microinjection studies in the Xenopus laevis oocyte, we have elucidated a novel pathway leading to the appearance of a processed B1-type Alu RNA species in the cytoplasm. The abundance of this small Alu RNA differs between various mouse tissues, suggesting a role in tissue-specific gene expression.

Animals↗
PubMed2004

Short interfering RNAs specific for potato spindle tuber viroid are found in the cytoplasm but not in the nucleus.

Short interfering (si) and micro (mi) RNAs influence gene expression at post-transcriptional level. In plants, different classes of DICER-LIKE (DCL) enzymes are responsible for the generation of these small regulatory RNAs from different precursors. To characterize the cellular site of their generation and accumulation, we purified nuclei from tomato plants infected with potato spindle tuber viroid (PSTVd) RNA, which is known to replicate in the nucleus via double-stranded (ds) RNA intermediates. We could detect PSTVd-specific siRNAs in the cytoplasmic fraction, but not in the nuclear fraction. To correlate the localization of the PSTVd-specific siRNAs with that of similarly sized small RNAs, we studied the compartmentalization of a naturally occurring miRNA. We could detect the precursor of miR167 in the nucleus, but the mature miRNA was found only in the cytoplasmic fraction. We discuss the consequences of this finding for the model of viroid replication and heterochromatin formation.

Cell Nucleus↗
PubMed1987

Small cytoplasmic RNAs and their location within the cytoplasm.

These studies were designed to establish the location of various species of small RNAs within the subcellular cytoplasmic compartments. Four cytoplasmic RNA-containing compartments were examined: (A) cytoskeleton-bound polyribosomal ribonucleoprotein (RNP) complexes, (B) soluble-phase polyribosomal RNP complexes, (C) cytoskeleton-bound free RNP complexes, (D) soluble-phase free RNP complexes. The presence of the small cytoplasmic RNA (scRNA) population and histone H4 and actin mRNAs in each compartment was examined to determine their spatial distribution within the cytoplasm. The 7S signal recognition RNA and the 5S and 5.8S rRNAs were distributed among all four compartments, while 4S tRNAs were localized largely in fraction D. Fraction C contained a group of seven abundant scRNAs, of approximately 105-348 nucleotides in length, which were localized almost entirely within the cytoskeleton-bound free RNP compartment. Actin mRNAs were localized in fraction A, the actively translating cytoskeleton-bound compartment. Actin mRNAs were localized in fraction A, the actively translating cytoskeleton-bound compartment. Following cytochalasin B treatment, actin mRNAs were released into the soluble phase, implicating a dependence on the integrity of actin filaments in its binding. Such treatment also released several of the scRNAs from their cytoskeleton-bound location. In contrast histone H4 mRNAs were much more widely dispersed, being present in all four cytoskeletal compartments. Approximately 60% of the H4 mRNAs, however, were localized within the soluble-phase polyribosomes in fraction B. Cytochalasin B treatment released only the small portion of untranslated histone H4 mRNA associated with the cytoskeleton in fraction C, suggesting that the binding of these H4 mRNAs was dependent in some manner upon the integrity of actin filaments.

Cytoplasm↗