Expression pattern of bta-mir-2898 miRNA and their correlation with heat shock proteins during summer heat stress among native vs crossbred cattle

Earlier studies identified the role of bta-mir-2898 in bovine. Our earlier study identified that, bta-mir-2898 can be over expressed in crossbred cattle during heat stress. Nevertheless the differential expression of bta-mir-2898 among native vs crossbred cattle during summer stress along with it's correlation with different heat shock proteins (HSPs) is not yet studied. In the present context, we studied the differential expression of bta-mir-2898 among Frieswal (Bos indicus x Bos taurus) and Sahiwal (Bos indicus) breeds of cattle during a range of environmental air temperatures and further investigated the correlation of bta-mir-2898 with different HSPs (HSP70, HSP90, HSP60. HSF, HSPB8 and HSP27). It was observed that, at peak air temperature the relative miRNA expression level (p < 0.05) of bta-mir-2898 was 3.4 ± 0.41 and 0.79 ± 0.22 among Frieswal and Sahiwal, respectively. We also observed significant levels (p < 0.05) of mRNA abundance of HSP70, HSP90, HSPB8 and HSP27 among the breeds. In all the cases Sahiwal found to exhibited higher level of HSPs in comparison to Frieswal. Studies revealed that the expression profile of bta-mir-2898 was negatively correlated with the expression of all the HSPs during thermal stress in post anti-mir2898 treated PBMC invitro cultured model originated from both Frieswal and Sahiwal cattle breeds. However, significantly (p < 0.05) higher negative correlations were observed between bta-mir-2898 and HSP70, HSP60 and HSPB8. Present findings highlighted the preliminary role of overexpressed bta-mir-2898 in cattle during thermal stress and its impact on different heat shock proteins.     

Identification and comparative analysis of the pearl oyster <Emphasis Type="Italic">Pinctada fucata</Emphasis> hemocytes microRNAs in response to <Emphasis Type="Italic">Vibrio alginolyticus</Emphasis> infection

MicroRNAs (miRNAs) are a class of noncoding RNA molecules that function as negative regulators of gene expression and play important roles in a wide spectrum of biological processes, including in immune response. However, the physiological regulation function of Pinctada fucata miRNAs, specially their immunomodulation has not been explored yet. Here, two small RNA libraries from hemocytes of P. fucata with or without Vibrio alginolyticus infection were constructed and sequenced using the high-throughput Illumina deep sequencing technology. In total, 11,939,992 and 11,083,327 raw reads, corresponding to 10,993,546 and 9,988,179 clean reads, were respectively obtained in the control and infected libraries. A total of 276 miRNAs, including 225 known miRNAs and 51 putative novel miRNAs, were identified by bioinformatic analysis. By using pairwise comparison between two libraries, 93 miRNAs were found to be significantly differentially expressed, with 42 and 51 miRNAs exhibiting up-regulation and down-regulation, respectively. Thereinto, some known miRNAs were considered to be immune-related. Real-time PCR were implemented for 6 miRNAs co-expressed in the control and infected samples, and agreement was confirmed between the high-throughput sequencing and real-time PCR data. After miRNA targets were predicted, GO and KEGG pathway enrichment analysis were performed, and the results indicated that ten of the differentially expressed miRNAs were involved in immune-related pathways, and might participate in the host immune response to V. alginolyticus. These results of identification and comparative analysis of miRNAs might deepen our understanding of host-pathogen interactions and immune defense mechanisms in P. fucata.     

Molecular cloning and characterization of the <Emphasis Type="Italic">MsHSP17.7</Emphasis> gene from <Emphasis Type="Italic">Medicago sativa</Emphasis> L.

Heat shock proteins (HSPs) are ubiquitous protective proteins that play crucial roles in plant development and adaptation to stress, and the aim of this study is to characterize the HSP gene in alfalfa. Here we isolated a small heat shock protein gene (MsHSP17.7) from alfalfa by homology-based cloning. MsHSP17.7 contains a 477-bp open reading frame and encodes a protein of 17.70-kDa. The amino acid sequence shares high identity with MtHSP (93.98 %), PsHSP17.1 (83.13 %), GmHSP17.9 (74.10 %) and SlHSP17.6 (79.25 %). Phylogenetic analysis revealed that MsHSP17.7 belongs to the group of cytosolic class II small heat shock proteins (sHSP), and likely localizes to the cytoplasm. Quantitative RT-PCR indicated that MsHSP17.7 was induced by heat shock, high salinity, peroxide and drought stress. Prokaryotic expression indicated that the salt and peroxide tolerance of Escherichia coli was remarkably enhanced. Transgenic Arabidopsis plants overexpressing MsHSP17.7 exhibited increased root length of transgenic Arabidopsis lines under salt stress compared to the wild-type line. The malondialdehyde (MDA) levels in the transgenic lines were significantly lower than in wild-type, although proline levels were similar between transgenic and wild-type lines. MsHSP17.7 was induced by heat shock, high salinity, oxidative stress and drought stress. Overexpression analysis suggests that MsHSP17.7 might play a key role in response to high salinity stress.     

<Emphasis Type="Italic">Hsp70</Emphasis> genes of the <Emphasis Type="Italic">Megaphragma amalphitanum</Emphasis> (Hymenoptera: Trichogrammatidae) parasitic wasp

Miniaturization is an evolutionary process that is widely represented in both invertebrates and vertebrates. Miniaturization frequently affects not only the size of the organism and its constituent cells, but also changes the genome structure and functioning. The structure of the main heat shock genes (hsp70 and hsp83) was studied in one of the smallest insects, the Megaphragma amalphitanum (Hymenoptera: Trichogrammatidae) parasitic wasp, which is comparable in size with unicellular organisms. An analysis of the sequenced genome has detected six genes that relate to the hsp70 family, some of which are apparently induced upon heat shock. Both induced and constitutively expressed hsp70 genes contain a large number of introns, which is not typical for the genes of this family. Moreover, none of the found genes form clusters, and they are all very heterogeneous (individual copies are only 75–85% identical), which indicates the absence of gene conversion, which provides the identity of genes of this family in Drosophila and other organisms. Two hsp83 genes, one of which contains an intron, have also been found in the M. amalphitanum genome.     

Revision of the genus <Emphasis Type="Italic">Melamphaes</Emphasis> (Melamphaidae). 5. Oligo-raker species: <Emphasis Type="Italic">M. indicus</Emphasis>, <Emphasis Type="Italic">M. eurous</Emphasis>, and <Emphasis Type="Italic">M. typhlops</Emphasis>

Three oligo-raker species (?19 rakers on the first gill arch) of the genus Melamphaes out of the “M. typhlops” group are considered. The validity of M. indicus Ebeling is restored. This species inhabits equatorial and tropical waters of the Indian Ocean and the western part of the Pacific Ocean. M. eurous sp. n., which is related to M. indicus, is described from equatorial waters of the eastern part of the Pacific Ocean. M. typhlops (Lowe) inhabiting the northern part of the Atlantic Ocean, from the equatorial zone about to 45° N, is redescribed.     

Calcium chloride improves photosynthesis and water status in the C<Subscript>4</Subscript> succulent xerophyte <Emphasis Type="Italic">Haloxylon ammodendron</Emphasis> under water deficit

It is important to reveal the mechanism of plants coping with heat stress, which results in a severe retardation in crop growth and development. Although Synaptotagmin A (SYTA) regulates the cell endocytosis and the plasma membrane reparation of Arabidopsis, its roles in heat resistance are not well understood. In this study, we obtained the new finding that SYTA is related to the heat resistance of Arabidopsis. In the SYTA overexpression strains, the germination rate of the seeds and the survival rate of the seedlings improved after heat shock treatment, and their degree of membrane peroxidation was reduced. However, syta mutant showed the opposite results. Meanwhile, the expressions of some heat stress signal pathway genes were higher in SYTA overexpression strains than that in wild-type strains, and were lower in syta mutant strains. These results suggested that SYTA responded positively to heat shock and was involved in the heat stress signal pathway.     

Over-Expression of <Emphasis Type="Italic">PmHSP17.9</Emphasis> in Transgenic <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> Confers Thermotolerance

Small heat shock proteins (sHSPs) have been shown to be involved in stress tolerance. However, their functions in Prunus mume under heat treatment are poorly characterized. To improve our understanding of sHSPs, we cloned a sHSP gene, PmHSP17.9, from P. mume. Sequence alignment and phylogenetic analysis indicated that PmHSP17.9 was a member of plant cytosolic class III sHSPs. Besides heat stress, PmHSP17.9 was also upregulated by salt, dehydration, oxidative stresses and ABA treatment. Leaves of transgenic Arabidopsis thaliana that ectopically express PmHSP17.9 accumulated less O₂ ^? and H₂O₂ compared with wild type (WT) after 42 °C treatment for 6 h. Over-expression of PmHSP17.9 in transgenic Arabidopsis enhanced seedling thermotolerance by decreased relative electrolyte leakage and MDA content under heat stress treatment when compared to WT plants. In addition, the induced expression of HSP101, HSFA2, and delta 1-pyrroline-5-carboxylate synthase (P5CS) under heat stress was more pronounced in transgenic plants than in WT plants. These results support the positive role of PmHSP17.9 in response to heat stress treatment.     

Small RNA changes in synthetic <Emphasis Type="Italic">Brassica napus</Emphasis>

Main conclusion

Small RNAs and microRNAs were found to vary extensively in synthetic Brassica napus and subsequent generations, accompanied by the activation of transposable elements in response to hybridization and polyploidization.

Abstract

Resynthesizing B. napus by hybridization and chromosome doubling provides an approach to create novel polyploids and increases the usable genetic variability in oilseed rape. Although many studies have shown that small RNAs (sRNAs) act as important factor during hybridization and polyploidization in plants, much less is known on how sRNAs change in synthetic B. napus, particularly in subsequent generations after formation. We performed high-throughput sequencing of sRNAs in S₁–S₄ generations of synthetic B. napus and in the homozygous B. oleracea and B. rapa parent lines. We found that the number of small RNAs (sRNAs) and microRNAs (miRNAs) doubled in synthetic B. napus relative to the parents. The proportions of common sRNAs detected varied from the S₁ to S₄ generations, suggesting sRNAs are unstable in synthetic B. napus. The majority of miRNAs (67.2 %) were non-additively expressed in the synthesized Brassica allotetraploid, and 33.3 % of miRNAs were novel in the resynthesized B. napus. The percentage of miRNAs derived from transposable elements (TEs) also increased, indicating transposon activation and increased transposon-associated miRNA production in response to hybridization and polyploidization. The number of target genes for each miRNA in the synthesized Brassica allotetraploid was doubled relative to the parents, enhancing the complexity of gene expression regulation. The potential roles of miRNAs and their targets are discussed. Our data demonstrate generational changes in sRNAs and miRNAs in synthesized B. napus.

     

Constitutive Expression of a Tomato Small Heat Shock Protein Gene <Emphasis Type="Italic">LeHSP21</Emphasis> Improves Tolerance to High-Temperature Stress by Enhancing Antioxidation Capacity in Tobacco

It is well established that small heat shock proteins (sHSPs) play an important role in thermotolerance in various organisms due to their abundance and diversity. In the present study, a chloroplast small heat shock protein gene (LeHSP21) from tomato (Lycopersicon esculentum cv PKM-1) was constitutively expressed in tobacco (Nicotiana tabacum L. cv Wisconsin 38) plants via Agrobacterium-mediated transformation. When compared to wild-type control plants, transgenic tobacco plants constitutively expressing LeHSP21, driven by the cauliflower mosaic virus 35S promoter, exhibited improved tolerance to both high temperature and oxidative stress. Furthermore, when the seedlings were subjected to high temperature treatment, the activities of anti-oxidative enzymes and the content of proline were significantly higher in transgenic plants than those in the wild-type plants. Our results presented here demonstrate the feasibility of improving high temperature and oxidative stress tolerance in plants through the expression of LeHSP21 gene.     

Ectopic Expression of Plant RNA Chaperone Offering Multiple Stress Tolerance in <Emphasis Type="Italic">E. coli</Emphasis>

Members of the plant glycine-rich RNA-binding proteins (GR-RBPs) family have been reported in flowering, development, circadian rhythms, biotic and abiotic stresses. Particularly, GR-RBPs are reported to function as RNA chaperones, promoting growth and acclimation during cold shock. It is indispensable to further question the efficacy and mechanism of GR-RBPs under various environmental strains. Monitoring the expression of stress-regulated proteins under stress conditions has been a beneficial strategy to study their functional roles. In an effort to elucidate the NtGR-RBP1 function, stress markers such as salinity, drought, low temperature and heat stresses were studied. The NtGR-RBP1 gene was expressed in E. coli followed by the exposure to stress conditions. Recombinant E. coli expressing NtGR-RBP1 were more tolerant to stresses, e.g., salinity, drought, cold and heat shock. Recombinants exhibited higher growth rates compared to control in spot assays. The tolerance was further confirmed by monitoring the growth in liquid culture assays. Cells expressing NtGR-RBP1 under salt (500 mM NaCl), drought (20% PEG), cold (4 and 20 °C) and heat stresses (50 °C) had enhanced growing ability and better endurance. Our study supports the notion that the protective role of NtGR-RBP1 may contribute to growth and survival during diverse environmental stresses.     

Integrating CNVs into meta-QTL identified <Emphasis Type="Italic">GBP4</Emphasis> as positional candidate for adult cattle stature

Copy number variation (CNV) of DNA sequences, functionally significant but yet fully ascertained, is believed to confer considerable increments in unexplained heritability of quantitative traits. Identification of phenotype-associated CNVs (paCNVs) therefore is a pressing need in CNV studies to speed up their exploitation in cattle breeding programs. Here, we provided a new avenue to achieve this goal that is to project the published CNV data onto meta-quantitative trait loci (meta-QTL) map which connects causal genes with phenotypes. Any CNVs overlapping meta-QTL therefore will be potential paCNVs. This study reported potential paCNVs in Bos taurus autosome 3 (BTA3). Notably, overview indexes and CNVs both highlighted a narrower region (BTA3 54,500,000–55,000,000 bp, named BTA3_INQTL_6) within one constructed meta-QTL. Then, we ascertained guanylate-binding protein 4 (GBP4) among the nine positional candidate genes was significantly associated with adult cattle stature, including body weight (BW, P?<?0.05) and withers height (WHT, P?<?0.05), fitting GBP4 CNV either with three levels or with six levels in the model. Although higher copy number downregulated the mRNA levels of GBP2 (P?<?0.05) and GBP4 (P?<?0.05) in 1-Mb window (54.0–55.0 Mb) in muscle and adipose, additional analyses will be needed to clarify the causality behind the ascertained association.