Search

The α subunit of avian myeloblastosis virus reverse transcriptase (AMV-RT) is generated from the β-subunit by proteolysis, and the αβ heterodimer represents the active form. The codon-optimized gene was expressed in Escherichia coli , and an active αβ heterodimer was generated. The RNA amplification activity of the purified recombinant AMV-RT αβ heterodimer was similar to that of the native one. [ABSTRACT FROM AUTHOR]

MYB (v-myb avian myeloblastosis viral oncogene homolog) transcription factors (TFs) participate in the abiotic stress response in several plant species. In this study, we cloned and characterised MdMYB44 from apple (Malus domestica). Phylogenetic analysis showed that the MdMYB44 protein was highly homologous to Arabidopsis thaliana MYB44. In addition, MdMYB44 localised to the nuclei in transformed onion epidermal cells and showed transcriptional activation activity in a yeast one-hybrid assay. MdMYB44 was expressed in the stem, leaves, roots, flowers, and fruits, with the highest transcript level being observed in the leaves. Quantitative PCR analysis demonstrated that MdMYB44 expression was down-regulated by cold, osmotic, and salt stresses. Furthermore, stable overexpression of MdMYB44 in transgenic apple calli remarkably alleviated their tolerance to high salinity and cold, and relative conductivity and malondialdehyde accumulation under salt and cold treatments were significantly higher than those in wild-type control calli. Therefore, our findings provide evidence that MdMYB44 is involved in susceptibility to cold and salt stress in apple calli and indicate MdMYB44 to negatively regulate plant abiotic stress defence. [ABSTRACT FROM AUTHOR]

The v-myb avian myeloblastosis viral oncogene homolog (MYB) superfamily constitutes one of the most abundant groups of transcription factors (TFs) described in plants. To date, little is known about the MYB genes in Rehmannia glutinosa. Forty unique MYB genes with full-length cDNA sequences were isolated. These 40 genes were grouped into five categories, one R1R2R3-MYB, four TRFL MYBs, four SMH MYBs, 25 R2R3-MYBs, and six MYB-related members. The MYB DNA-binding domain (DBD) sequence composition was conserved among proteins of the same subgroup. As expected, most of the closely related members in the phylogenetic tree exhibited common motifs. Additionally, the gene structure and motifs of the R. glutinosa MYB genes were analyzed. MYB gene expression was analyzed in the leaf and the tuberous root under two abiotic stress conditions. Expression profiles showed that most R. glutinosa MYB genes were expressed in the leaf and the tuberous root, suggesting that MYB genes are involved in various physiological and developmental processes in R. glutinosa. Seven MYB genes were up-regulated in response to shading in at least one tissue. Two MYB genes showed increased expression and 13 MYB genes showed decreased expression in the tuberous root under continuous cropping. This investigation is the first comprehensive study of the MYB gene family in R. glutinosa. [ABSTRACT FROM AUTHOR]

Background Formation of a mature core is a crucial event for infectivity of retroviruses such as Mason- Pfizer monkey virus (M-PMV). The process is triggered by proteolytic cleavage of the polyprotein precursor Gag, which releases matrix, capsid (CA), and nucleocapsid proteins. Once released, CA assembles to form a mature core - a hexameric lattice protein shell that protects retroviral genomic RNA. Subtle conformational changes within CA induce the transition from the immature lattice to the mature lattice. Upon release from the precursor, the initially unstructured N-terminus of CA is refolded to form a β-hairpin stabilized by a salt bridge between the N-terminal proline and conserved aspartate. Although the crucial role of the β-hairpin in the mature core assembly has been confirmed, its precise structural function remains poorly understood. Results Based on a previous NMR analysis of the N-terminal part of M-PMV CA, which suggested the role of additional interactions besides the proline-aspartate salt bridge in stabilization of the β-hairpin, we introduced a series of mutations into the CA sequence. The effect of the mutations on virus assembly and infectivity was analyzed. In addition, the structural consequences of selected mutations were determined by NMR spectroscopy. We identified a network of interactions critical for proper formation of the M-PMV core. This network involves residue R14, located in the N-terminal β-hairpin; residue W52 in the loop connecting helices 2 and 3; and residues Q113, Q115, and Y116 in helix 5. Conclusion Combining functional and structural analyses, we identified a network of supportive interactions that stabilize the β-hairpin in mature M-PMV CA. [ABSTRACT FROM AUTHOR]

V-myb avian myeloblastosis viral oncogene homolog (MYB) transcription factors play important roles in the processes of plant growth, development, and stress responses. In this study, a full-length cDNA sequence of a MYB gene, designated FvMYB1, was isolated from Arizona ash (Fraxinus velutina Torr.) for the first time. Sequence analysis showed that the deduced amino acid sequence of FvMYB1 encoded novel MYB proteins with single DNA binding domains. Based on the R3 domain amino acid sequence, the FvMYB1 was closely related to some proteins whose functions were known. The expression pattern of FvMYB1 in different organs of Arizona ash was analyzed by semiquantitative RT-PCR and real-time PCR (qRT-PCR). Results showed that this gene was widely distributed in all the tested organs and the expression level of FvMYB1 was the highest in stems and the lowest in roots. The gene expression level decreased dramatically in roots under salt treatment (300 mM, 24 h), while no obvious change was observed in stems. Subcellular localization indicated that FvMYB1 was localized in the nucleus. This study will lead to further research in resistance to abiotic stress in Arizona ash. [ABSTRACT FROM AUTHOR]

The Myb protein was first identified as an oncogene that causes leukemia in chickens. Since then, it has been widely associated with different types of cancers and studied in detail in myeloid leukemias. However, despite these studies, its role in the induction, pathogenesis and maintenance of AML, and other blood disorders, is still not well understood. Recent efforts to uncover its plethora of transcriptional targets have provided key insights into understanding its mechanism of action. This review evaluates our current knowledge of the role of Myb in leukemia, with a particular focus on AML, from the vast literature spanning three decades, highlighting key studies that have influenced our understanding. We discuss recent insights into its role in leukemogenesis and how these could be exploited for the therapeutic targeting of Myb, its associated co-regulators or its target genes, in order to improve outcomes in the treatment of a wide range of hematopoietic malignancies. [ABSTRACT FROM AUTHOR]

Abstract: We investigated the effects of fibroblast growth factor-21 (FGF-21) on palmitate-induced insulin resistance in skeletal muscle myotubes. First, to determine the effect of FGF-21 on palmitate-induced insulin resistance, we measured 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-d-glucose uptake and levels of proteins involved in insulin signaling pathways (IRS-1 and Akt) in human skeletal muscle myotubes (HSMMs) exposed to palmitate for 24h, and compared HSMMs exposed to palmitate and different doses of recombinant FGF-21. Second, to determine the mechanisms underlying the contribution of FGF-21 to palmitate-induced insulin resistance, we compared levels of proteins linked to palmitate-induced insulin resistance (PKC-θ, IKKα/β, JNK, p38, IκBα, and NF-κB) in HSMMs exposed to palmitate and different doses of recombinant FGF-21 for 24h. Palmitate-reduced glucose uptake was restored by FGF-21. Palmitate inhibited phosphorylation of Akt and thereby impaired insulin signaling in HSMMs. FGF-21 prevented palmitate from inhibiting the phosphorylation of Akt. These results indicate that FGF-21 prevented palmitate-induced insulin resistance in HSMMs. Palmitate activated NF-κB in HSMMs, thereby impairing the action of insulin and initiating chronic inflammation. FGF-21 inhibited palmitate-induced NF-κB activation in HSMMs. The results of the present study suggest that FGF-21 prevents palmitate-induced insulin resistance in HSMMs by inhibiting the activation of stress kinase and NF-κB. [Copyright &y& Elsevier]

Avian myeloblastosis virus reverse transcriptase (AMV RT) is a heterodimer consisting of a 63 kDa α-subunit and a 95 kDa β subunit. Moloney murine leukaemia virus reverse transcriptase (MMLV RT) is a 75 kDa monomer. These two RTs are the most extensively used for conversion of RNA to DNA. We previously developed several mutations that increase the thermostability of MMLV RT and generated a highly stable MMLV RT variant E286R/E302K/L435R/D524A by combining three of them (Glu286→Arg, Glu302→Lys, and Leu435→Arg) and the mutation to abolish RNase H activity (Asp524→Ala) [Yasukawa et al. (2010) J Biotechnol 150:299-306]. To generate a highly stable AMV RT variant, we have introduced the triple mutation of Val238→Arg, Leu388→Arg, and Asp450→Ala into AMV RT α-subunit and the resulted variant V238R/L388R/D450A, was expressed in insect cells and purified. The temperature decreasing the initial activity by 50 %, measured over 10 min, of the variant with or without template primer (T/P), poly(rA)-p(dT), was 50 °C; for the wild-type AMV RT α-subunit (WT) this was 44 °C. The highest temperature at which the variant exhibited cDNA synthesis activity was 64 °C; the WT was 60 °C. A highly stable AMV RT α-subunit is therefore generated by the same mutation strategy as applied to MMLV RT and that positive charges are introduced into RT at positions that have been implicated to interact with T/P by site-directed mutagenesis. [ABSTRACT FROM AUTHOR]

Abstract: The epididymis plays a crucial role in regulating the development of sperm motility and fertilizing capacity. Small non-coding RNAs (sncRNAs), especially microRNAs (miRNAs), can participate in the regulation of various physiological pathways. However, their abundance and whether they are involved in the regulation of gene expression in the human epididymis are unknown. By adopting the Solexa deep sequencing approach, we systematically investigated the sncRNAs in the adult human epididymis. A total of 4903 unique sequences representing 527 known miRNA were discovered. Eighteen novel miRNA genes encoding 23 mature miRNAs were also identified and the expression of some of them was confirmed by qRT-PCR. The presence of Piwi-interacting RNAs (piRNAs) in the library also adds to the diversity of the sncRNA population in the human epididymis. This research will contribute to a preliminary database for their functional study in male reproductive system. [Copyright &y& Elsevier]

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

The article offers information on Desert hedgehog (DHH) which belongs to hedgehog gene family. It states that DHH is an essential morphogen for normal testicular development. It mentions that DHH is presents in mice and humans but not in avian lineage. It also determines that DHH signalling is an essential factor in gonadal development and gametogenesis.

Reverse transcriptases (RTs) from Moloney murine leukaemia virus (MMLV) and avian myeloblastosis virus (AMV) contain all the fingers, palm, thumb, connection and RNase H domains. The fingers, palm and thumb domains are thought to be involved in the reverse transcription activity, and the RNase H domain is in the RNase H activity. In this study, we characterized four chimeric RTs which comprise one of the fingers, palm, thumb and RNase H domains originated from AMV RT and the other three and connection domains originated from MMLV RT. Unexpectedly, all chimeric RTs exhibited the same characteristics: their specific reverse transcription activities decreased to less than 0.1% of that of MMLV RT, while their specific RNase H activities were approximately 20% of that of MMLV RT. The decreases in the two activities of the chimeric RTs were ascribed to the decreases in kcat. Based on that the reverse transcription activity of MMLV RT was impaired by substituting its RNase H domain with that from AMV RT, we propose that in MMLV RT, there might be an interaction between the fingers/palm/thumb domain and the RNase H domain. [ABSTRACT FROM PUBLISHER]

Reverse transcriptases (RTs) from avian myeloblastosis virus (AMV) and Moloney murine leukaemia virus (MMLV) have been most extensively used as a tool for conversion of RNA to DNA. In this study, we compared the thermal stabilities of AMV RT and MMLV RT by observing their irreversible thermal inactivation. The temperatures reducing initial activity by 50% in 10-min incubation, T50, of AMV RT were 47°C without the template-primer (T/P), poly(rA)–p(dT)12–18, and 52°C with the T/P (28 μM). T50 of MMLV RT were 44°C without the T/P and 47°C with the T/P (28 μM). Unexpectedly, AMV RT was considerably activated when incubated with the T/P at 45 and 48°C. Such activation was not observed in MMLV RT. These results suggest that AMV RT and MMLV RT are different in the following: (i) The intrinsic thermal stability of AMV RT is higher than that of MMLV RT; (ii) AMV RT is activated by thermal treatment with the T/P at 45–48°C; and (iii) AMV RT is stabilized by the T/P more potently than MMLV RT. Thermodynamic analysis indicates that thermal inactivation of AMV RT and MMLV RT is due to the large entropy change of activation for thermal inactivation. [ABSTRACT FROM PUBLISHER]

A TaqMan fluorescent probe-based product enhanced reverse transcriptase (RT) assay is described in which the RT and polymerase chain reaction (PCR) steps are set-up in a single tube, in two compartments separated by Ampliwax™ (designated as single-tube fluorescent product-enhanced reverse transcriptase assay (STF-PERT)). This simplification of the two-step method resulted in increased assay reproducibility and handling efficiency while maintaining the sensitivity of the PERT assay (<10 virions). The STF-PERT assay can be used to quantitate low amounts of retrovirus in clinical and research materials and to evaluate retrovirus contamination in cell substrates and biological products in human use. [Copyright &y& Elsevier]

A formulation of reverse transcriptases comprising enzymes derived from avian myeloblastosis virus and from Moloney murine leukaemia virus has provided an improved method for amplifying nucleic acid sequences by coupled reverse transcription and polymerase chain reaction. The properties of each enzyme appear to complement the other and demonstrate enhanced sensitivity and increased product yield in comparison to either enzyme used separately. [ABSTRACT FROM AUTHOR]

We have recently expressed in bacteria an enzymatically active reverse transcriptase (RT) of mouse mammary tumor virus (MMTV), a mammalian retrovirus with a typical B-type morphology [Taube, R., Loya, S., Avidan, O., Perach, M. & Hizi, A. (1998) Biochemical J. 329, 579-587]. The purified recombinant protein was shown to possess the catalytic activities characteristic of retroviral reverse transcriptases. In the present study, we have analyzed two basic parameters characteristic of the DNA polymerase activity of the novel MMTV RT, namely the processivity and the fidelity of DNA synthesis. Two features related to fidelity were studied, the capacity to misinsert wrong nucleotides at the 3′ end of the nascent DNA strand and the ability to extend 3′ mispairs. The studied properties of MMTV RT were compared with those of the RT purified from virions of avian myeloblastosis virus (AMV), since AMV RT shows a relatively high sequence similarity to MMTV RT. MMTV RT shows a relative processivity of DNA synthesis which is as high as the reference AMV RT. Regarding fidelity of DNA synthesis, MMTV RT shows a fidelity of misinsertion lower than that of AMV RT, whereas its capacity to elongate mispaired DNA is lower than that of AMV RT indicating a somewhat higher fidelity. These fidelity properties are discussed also in the context of the RTs of lentiviruses, especially those of HIV, which were reported to exhibit an exceptionally low fidelity of DNA synthesis. It is clear that MMTV RT has a fidelity higher than that of lentiviral RTs. [ABSTRACT FROM AUTHOR]

An aqueous extract from the marine red alga, Schizymenia pacifica has been tested in a cell free system for its effect on reverse transcriptase from avian retrovirus (avian myeloblastosis virus), and mammalian retrovirus (Rauscher murine leukemia virus). The extract inhibited reverse transcriptase from both these retroviruses but showed almost no effect, if any, on the activity of cellular DNA polymerase alpha and RNA polymerase II in vitro. Consequently it is unlikely to have an adverse effect on the growth of cultured cell. The inhibitory activity of the extract was stable over a relatively wide pH range (pH 1-11) and was not lost after pronase digestion. Inhibitory activity of the extract was lost after boiling at 100°C in 0.67 N HCl, and after treatment with 100 m M NaIO. The active principle in the extract has an apparent molecular weight in excess of 100 000 daltons. This new reverse transcriptase inhibitor is probably a polysaccharide. [ABSTRACT FROM AUTHOR]