Structure, Evolution, and Mitochondrial Genome Analysis of Mussel Species (Bivalvia, Mytilidae).

Based on the nucleotide sequences of the mitochondrial genome (mitogenome) of specimens taken from two mussel species (Arcuatula senhousia and Mytilus coruscus), an investigation was performed by means of the complex approaches of the genomics, molecular phylogenetics, and evolutionary genetics. The mitogenome structure of studied mussels, like in many other invertebrates, appears to be much more variable than in vertebrates and includes changing gene order, duplications, and deletions, which were most frequent for tRNA genes; the mussel species' mitogenomes also have variable sizes. The results demonstrate some of the very important properties of protein polypeptides, such as hydrophobicity and its determination by the purine and pyrimidine nucleotide ratio. This fact might indirectly indicate the necessity of purifying natural selection for the support of polypeptide functionality. However, in accordance with the widely accepted and logical concept of natural cutoff selection for organisms living in nature, which explains its action against deleterious nucleotide substitutions in the nonsynonymous codons (mutations) and its holding of the active (effective) macromolecules of the polypeptides in a population, we were unable to get unambiguous evidence in favor of this concept in the current paper. Here, the phylogeny and systematics of mussel species from one of the largest taxons of bivalve mollusks are studied, the family known as Mytilidae. The phylogeny for Mytilidae (order Mytilida), which currently has no consensus in terms of systematics, is reconstructed using a data matrix of 26-27 mitogenomes. Initially, a set of 100 sequences from GenBank were downloaded and checked for their gender: whether they were female (F) or male (M) in origin. Our analysis of the new data confirms the known drastic differences between the F/M mitogenome lines in mussels. Phylogenetic reconstructions of the F-lines were performed using the combined set of genetic markers, reconstructing only protein-coding genes (PCGs), only rRNA + tRNA genes, and all genes. Additionally, the analysis includes the usage of nucleotide sequences composed of other data matrices, such as 20-68 mitogenome sequences. The time of divergence from MRCA, estimated via BEAST2, for Mytilidae is close to 293 Mya, suggesting that they originate in the Silurian Period. From all these data, a consensus for the phylogeny of the subfamily of Mytilinae and its systematics is suggested. In particular, the long-debated argument on mussel systematics was resolved as to whether Mytilidae, and the subfamily of Mytilinae, are monophyletic. The topology signal, which was strongly resolved in this paper and in the literature, has refuted the theory regarding the monophyly of Mytilinae.

Long Noncoding RNAs in Response to Hyperosmolarity Stress, but Not Salt Stress, Were Mainly Enriched in the Rice Roots.

Due to their immobility and possession of underground parts, plants have evolved various mechanisms to endure and adapt to abiotic stresses such as extreme temperatures, drought, and salinity. However, the contribution of long noncoding RNAs (lncRNAs) to different abiotic stresses and distinct rice seedling parts remains largely uncharacterized beyond the protein-coding gene (PCG) layer. Using transcriptomics and bioinformatics methods, we systematically identified lncRNAs and characterized their expression patterns in the roots and shoots of wild type (WT) and ososca1.1 (reduced hyperosmolality-induced [Ca2+]i increase in rice) seedlings under hyperosmolarity and salt stresses. Here, 2937 candidate lncRNAs were identified in rice seedlings, with intergenic lncRNAs representing the largest category. Although the detectable sequence conservation of lncRNAs was low, we observed that lncRNAs had more orthologs within the Oryza. By comparing WT and ososca1.1, the transcription level of OsOSCA1.1-related lncRNAs in roots was greatly enhanced in the face of hyperosmolality stress. Regarding regulation mode, the co-expression network revealed connections between trans-regulated lncRNAs and their target PCGs related to OsOSCA1.1 and its mediation of hyperosmolality stress sensing. Interestingly, compared to PCGs, the expression of lncRNAs in roots was more sensitive to hyperosmolarity stress than to salt stress. Furthermore, OsOSCA1.1-related hyperosmolarity stress-responsive lncRNAs were enriched in roots, and their potential cis-regulated genes were associated with transcriptional regulation and signaling transduction. Not to be ignored, we identified a motif-conserved and hyperosmolarity stress-activated lncRNA gene (OSlncRNA), speculating on its origin and evolutionary history in Oryza. In summary, we provide a global perspective and a lncRNA resource to understand hyperosmolality stress sensing in rice roots, which helps to decode the complex molecular networks involved in plant sensing and adaptation to stressful environments.

Establishment and Maintenance of Human CRC-Derived Organoids for PcG Studies.

In the recent years, the establishment of self-organizing 3D cultures (organoids) generated from human primary tissues added a novel and physiological viewpoint to interrogate basic and pathological matters. Indeed, these 3D mini-organs, contrary to cell lines, faithfully reproduce the architecture and the molecular features of their original tissues. In cancer studies, the use of tumor patient-derived organoids (PDOs), capturing the histological and molecular heterogeneity of "pure" cancer cells, offered the opportunity to deeply explore tumor-specific regulatory networks. Accordingly, the study of polycomb group proteins (PcGs) can take advantage from this versatile technology to thoroughly investigate the molecular activity of these master regulators. In particular, the application of chromatin immunoprecipitations (ChIP)-sequencing (ChIP-seq) analyses to organoid models provides a powerful tool toward an accurate inquiry of PcG role in tumor development and maintenance.

The mitochondrial genomes of big-eared bats, Macrotus waterhousii and Macrotus californicus (Chiroptera: Phyllostomidae: Macrotinae).

In the species-rich family Phyllostomidae, the genus Macrotus ('big eared' bats) contains only two species; Macrotus waterhousii, distributed in western, central, and southern Mexico, Guatemala and some Caribbean Islands, and Macrotus californicus, distributed in the southwestern USA, and in the Baja California peninsula and the state of Sonora in Mexico. In this study, we sequenced and assembled the mitochondrial genome of Macrotus waterhousii and characterized in detail this genome and that of the congeneric M. californicus. Then, we examined the phylogenetic position of Macrotus in the family Phyllostomidae based on protein coding genes (PCGs). The AT-rich mitochondrial genomes of M. waterhousii and M. californicus are 16,792 and 16,691 bp long, respectively, and each encode 13 PCGs, 22 tRNA genes, 2 rRNA genes, and a putative non-coding control region 1,336 and 1,232 bp long, respectively. Mitochondrial synteny in Macrotus is identical to that reported before for all other cofamilial species. In the two studied species, all tRNAs exhibit a 'typical' cloverleaf secondary structure with the exception of trnS1, which lacks the D arm. A selective pressure analysis demonstrated that all PCGs are under purifying selection. The CR of the two species feature three domains previously reported in other mammals, including bats: extended terminal associated sequences (ETAS), central (CD), and conserved sequence block (CSB). A phylogenetic analysis based on the 13 mitochondrial PCGs demonstrated that Macrotus is monophyletic and the subfamily Macrotinae is a sister group of all remaining phyllostomids in our analysis, except Micronycterinae. The assembly and detailed analysis of these mitochondrial genomes represents a step further to continue improving the understanding of phylogenetic relationships within the species-rich family Phyllostomidae.

Cistanche Species Mitogenomes Suggest Diversity and Complexity in Lamiales-Order Mitogenomes.

The extreme diversity and complexity of angiosperms is well known. Despite the fact that parasitic plants are angiosperms, little is known about parasitic plant mitogenomic diversity, complexity, and evolution. In this study, we obtained and characterized the mitogenomes of three Cistanche species (holoparasitic plants) from China to compare the repeats, segment duplication and multi-copy protein-coding genes (PCGs), to clarify the phylogenetic and evolution relationship within the Lamiales order, and to identify the mitochondrial plastid insertions (MTPT) in Cistanche mitogenomes. The results showed that the mitogenome sizes of the three Cistanche species ranged from 1,708,661 to 3,978,341 bp. The Cistanche species genome encodes 75-126 genes, including 37-65 PCGs, 31-58 tRNA genes and 3-5 rRNA genes. Compared with other Lamiales and parasitic species, the Cistanche species showed extremely high rates of multi-copy PCGs, ranging from 0.13 to 0.58 percent of the total number of PCGs. In addition, 37-133 Simple Sequence Repeat (SSRs) were found in these three mitogenomes, the majority of which were the mononucleotides Adenine/Thymine. The interspersed repeats contained forward and palindromic repeats. Furthermore, the segment-duplication sequence size ranged from 199,584 to 2,142,551 bp, accounting for 24.9%, 11.7% and 53.9% of the Cistanche deserticola, Cistanche salsa and Cistanche tubulosa mitogenome, respectively. Furthermore, the Ka/Ks analysis suggested that the atp4, ccmB, ccmFc and matR were probably positively selected during Lamiales evolution. The Cistanche plastome suggested the presence of MTPT. Moreover, 6-12 tRNA, 9-15 PCGs fragments and 3 rRNA gene fragments in the Cistanche mitogenomes were found in the MTPT regions. This work reports the Cistanche species mitogenome for the first time, which will be invaluable for study the mitogenome evolution of Orobanchaceae family.

CirRNAPL: A web server for the identification of circRNA based on extreme learning machine.

Circular RNA (circRNA) plays an important role in the development of diseases, and it provides a novel idea for drug development. Accurate identification of circRNAs is important for a deeper understanding of their functions. In this study, we developed a new classifier, CirRNAPL, which extracts the features of nucleic acid composition and structure of the circRNA sequence and optimizes the extreme learning machine based on the particle swarm optimization algorithm. We compared CirRNAPL with existing methods, including blast, on three datasets and found CirRNAPL significantly improved the identification accuracy for the three datasets, with accuracies of 0.815, 0.802, and 0.782, respectively. Additionally, we performed sequence alignment on 564 sequences of the independent detection set of the third data set and analyzed the expression level of circRNAs. Results showed the expression level of the sequence is positively correlated with the abundance. A user-friendly CirRNAPL web server is freely available at http://server.malab.cn/CirRNAPL/.

The WNT Pathway Is Relevant for the BCR-ABL1-Independent Resistance in Chronic Myeloid Leukemia.

Notwithstanding the introduction of Tyrosine Kinase Inhibitors (TKIs) revolutionized the outcome of Chronic Myeloid Leukemia (CML), one third of patients still suspends treatment for failure response. Recent research demonstrated that several BCR/ABL1-independent mechanisms can sustain resistance, but the relationship between these mechanisms and the outcome has not yet been fully understood. This study was designed to evaluate in a "real-life" setting if a change of expression of several genes involved in the WNT/BETA-CATENIN, JAK-STAT, and POLYCOMB pathways might condition the outcome of CML patients receiving TKIs. Thus, the expression of 255 genes, related to the aforementioned pathways, was measured by quantitative PCR after 6 months of therapy and compared with levels observed at diagnosis in 11 CML patients, in order to find possible correlations with quality of response to treatment and event-free-survival (EFS). These results were then re-analyzed by the principal component method (PCA) for tempting to better cluster resistant cases. After 12 months of therapy, 6 patients achieved an optimal response and 5 were "resistant;" after application of both statistical methods, it was evident that in all pathways a significant overall up-regulation occurred, and that WNT was the pathway mostly responsible for the TKIs resistance. Indeed, 100% of patients with a "low" up-regulation of this pathway achieved an optimal response vs. 33% of those who showed a "high" gene over-expression (p = 0.016). Analogously, the 24-months EFS resulted significantly influenced by the degree of up-regulation of the WNT signaling: all patients with a "low" up-regulation were event-free vs. 33% of those who presented a "high" gene expression (p = 0.05). In particular, the PCA analysis confirmed the role of WNT pathway and showed that the most significantly up-regulated genes with negative prognostic value were DKK, WNT6, WISP1, and FZD8. In conclusion, our results sustain the need of a wide and multitasking approach in order to understand the resistance mechanisms in CML.

The 2018 risk-adjustment reform in the Czech Republic: Introducing Pharmacy-based Cost Groups and strengthening reinsurance.

BACKGROUND: Risk-adjustment in resource allocation is commonly used for regional redistribution or for eliminating risk selection motives of multiple statutory health insurers. In the Czech Republic, revenue redistribution between health insurers takes place since the 1990's. Since 2018, the risk-adjustment mechanism includes an adjustment for insured with chronic diseases using Pharmacy-based Cost Group (PCG) classification. In addition, retrospective compensation for very high cost patients has been strengthened. AIM: To provide an internationally relevant overview of the Czech risk-adjustment system. To assess the implication of the 2018 reform for health insurers and for the development of chronic care. METHOD: The framework of the Health Reform Monitor is used to analyse the policy process. Data from Czech health insurers and Czech Ministry of Health are used to assess likely impact of the reform. RESULTS: The reform increases coverage of predictable individual health risks and combines prospective risk-rating with strengthened retrospective risk-sharing among insurers. The reform results in moderate changes in risk-adjusted allocations of individual insurers. CONCLUSION: The Czech experience with risk-adjustment reforms is relevant for countries with multiple health insurers as well as for countries with risk-adjusted regional redistribution mechanisms. Combining prospective risk factors of age, sex, and PCGs with retrospective compensation of expensive cases limits potential losses to a manageable level, also for small risk-pools. It reduces incentives for cream skimming based on health status, enables higher use of risk-sharing contracts, and incentivizes the development of disease management programs in the Czech Republic.

The complete mitochondrial genome of a tropical sea cucumber, Holothuria leucospilota.

The mitochondrial genome of Holothuria leucospilota was 15,906 bp in length, containing 13 protein-coding genes (PCGs), 22 tRNA genes, and two rRNA genes. There were four initiation codons (ATG, ATT, ATC, and ATA) for the PCGs, and the termination codon of most PCGs was TAA, except for nad4 (TAG) and nad6 (TAG). Only one PCG (nad6) and five tRNA genes (tRNASer(UCN) , tRNAGln , tRNAAla , RNAVal , and tRNAAsp ) were encoded on the light chain; the other genes were encoded on the heavy chain. H. leucospilota was most closely related to Holothuria scabra in a phylogenetic tree.

Complete mitochondrial genome of the stonefly Cryptoperla stilifera Sivec (Plecoptera: Peltoperlidae) and the phylogeny of Polyneopteran insects.

We present the complete mitogenome of a stonefly, Cryptoperla stilifera Sivec (Plecoptera; Peltoperlidae). The mitogenome was a circular molecule consisting of 15,633 nucleotides, 37 genes and a A+T-rich region. C. stilifera mitogenome was similar to Pteronarcys princeps mitogenome (Plecoptera; Pteronarcyidae). All transfer RNA genes (tRNAs) had typical cloverleaf secondary structures except for trnSer (AGN), where the stem-loop structure of the dihydrouridine (DHU) arm was missing. The A+T-rich region of C. stilifera had two stem-loops and each had two interlink. Three conserved sequence blocks (CSBs) were present in the A+T-rich regions of C. stilifera, Peltoperla tarteri and Peltoperla arcuata. Moreover, many polynucleotide stretches (Poly N, N=A, T and C) in the A+T-rich region of C. stilifera Phylogenetic relationships of Polyneopteran species were constructed based on the nucleotide sequences of 13 protein coding genes (PCGs). Both maximum likelihood (ML) and Bayesian inference (BI) analyses supported Grylloblattodea as the sister group to Plecoptera+Dermaptera and Embiidina and Phasmatodea as sister groups.

The complete mitochondrial genome sequence of Sogatella furcifera (Horváth) and a comparative mitogenomic analysis of three predominant rice planthoppers.

The white-backed planthopper (WBPH), Sogatella furcifera (Horváth), is one of the most destructive pests of rice crops in many Asian countries. Using long-PCR and shotgun library methods, we sequenced the entire mitochondrial genomes (mt-genomes) of two WBPH individuals. Total lengths of the mt-genome of the two WBPH individuals were 16,612 bp and 16,654 bp with an identical AT content of 76.19%. Among the 13 protein coding genes (PCGs), only nad5 used an atypical initiation codon GTG. Most of the tRNA genes had the typical cloverleaf secondary structure except that the dihydrouridine (DHU) arms in two trnS genes and the TΨC arm of trnG gene did not form a stable stem-loop structure. Similar to the brown planthopper (BPH), Nilaparvata lugens (Stål), and the small brown planthopper (SBPH), Laodelphax striatellus (Fallén), some extraordinary features were observed in the WBPH mt-genome. These include similar gene rearrangement pattern, unusually short length of the atp8 gene and variable numbers of tandem repeat (VNTR) structure in control region. Interestingly, the same tandem repeat unit with stable secondary structure appeared in two different planthoppers, WBPH and SBPH, which belong to two different genera of the Delphacidae. This peculiar feature provides a direct evidence for the close relationship between the two planthoppers and updates our understanding of the evolutionary characteristics of mitochondrial control region. Comparison with two other predominant rice planthoppers (BPH and SBPH) revealed that different PCGs of mitochondria exhibit different evolutionary patterns.

Identification of abiotic stress miRNA transcription factor binding motifs (TFBMs) in rice.

Plant growth and yield are affected by many abiotic stresses like salinity, drought, cold and heavy metal; these stresses trigger up and down-regulate several genes through various transcription factors (TFs). Transcription factor binding motifs (TFBMs), located in the upstream region of the genes, associate with TFs to regulate the gene expression. Many factors, including the activation of miRNAs, which are encoded by genes having independent transcription units, regulate the gene expression. TFBMs in the regulatory region of miRNA sequences influence the miRNA expression, which in turn influences the expression of other genes in the cell. However, the current level of information available on TFBMs of miRNA involved in abiotic stress related defense pathway(s) is limited and in-depth studies in this direction may lead to a better understanding of their role in expression and regulation of defense responses in plants. In this study, various aspects related to genomic positions of pre-miRNA, prediction of TSS and TATA box positions and identification of known, unique motifs at regulatory regions of all the reported miRNAs of rice associated with different abiotic stresses are discussed. Sixteen motifs were identified in this study, of which nine are known cis-regulatory elements associated with various stresses, two strong motifs, (CGCCGCCG, CGGCGGCG) and five unique motifs which might play a vital role in the regulation of abiotic stresses related miRNA genes. Common motifs shared by miRNAs that are involved in more than one abiotic stresses were also identified. The motifs identified in this study will be a resource for further functional validation.

The complete mitochondrial genome of the sycamore lace bug Corythucha ciliata (Hemiptera: Tingidae).

The complete mitochondrial genome of the sycamore lace bug, Corythucha ciliata, was sequenced in this study. It represents the first sequenced mitogenome of family Tingidae in Heteroptera. The mitogenome of C. ciliata is 15,257bp and contains 37 genes including 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes and a large non-coding region. Gene arrangement, nucleotide content, codon usage, and amino acid composition and asymmetry indicate a high degree of conservation with six other species of Cimicomorpha. The 13 PCGs initiated with ATN as the start codon and terminated with TAA, TA or T as stop codon. The evolutionary rate of each PCG was different, among which ATP8 showed the highest rate while ATP6 indicated the lowest rate. The 22 tRNAs genes apparently fold into a typical cloverleaf structure; however, the anticodon (TTC) of trnSer (AGN) differs from other Heteropteran insects. Secondary structure modeling of rRNA genes revealed similarity to other insects, except for two incomplete helices (H1648 and H2735) in lrRNA. The predicted secondary structure of lrRNA indicates 45 helices in six domains, whereas srRNA has 27 helices in three domains. Three potential stem-loops and two tandem repeats (-TCTAAT-) were identified in the A+T-rich region. Phylogenetic analysis indicated that C. ciliata is a sister group to other Heteroptera species based on analysis of the 13 PCGs.

The complete mitochondrial genome of the Ailanthus silkmoth, Samia cynthia cynthia (Lepidoptera: Saturniidae).

The complete mitochondrial genome (mitogenome) of the Ailanthus silkmoth, Samia cynthia cynthia (Lepidoptera: Saturniidae) was determined. The circular genome is 15,345 bp long, and presents a typical gene organization and order for sequenced mitogenomes of Bombycidea species. The nucleotide composition of the genome is highly A+T biased, accounting for 79.86%. The AT skew of the genome is slightly negative, indicating the occurrence of more Ts than As, as found in other Saturniidae species. All protein-coding genes (PCGs) are initiated by ATN codons, except for COI and COII, which are tentatively designated by CGA and GTG, respectively, as observed in other insects. Four of 13 PCGs, including COI, COII, ATP6, and ND3, harbor the incomplete termination codons, T or TA. With an exception for tRNASer(AGN), all other tRNAs can form a typical clover-leaf structure of mitochondrial tRNA. The 359 bp A+T-rich region of S. c. cynthia contains non-repetitive sequences, but harbors several features common to the Bombycidea insects, including the motif ATAGA followed by a poly-T stretch of 19 bp, a microsatellite-like (AT)7 element preceded by the ATTTA motif, and a poly-A element upstream tRNAMet. The phylogenetic analyses support the morphology-based current hypothesis that Bombycidae and Saturniidae are monophyletic. Our result confirms that Saturniini and Attacini form a reciprocal monophyletic group within Saturniidae.