LncRNA 4930581F22Rik promotes myogenic differentiation by regulating the ERK/MAPK signaling pathway.

The skeletal muscle is the largest organ in mammals and is the primary motor function organ of the body. Our previous research has shown that long non-coding RNAs (lncRNAs) are significant in the epigenetic control of skeletal muscle development. Here, we observed progressive upregulation of lncRNA 4930581F22Rik expression during skeletal muscle differentiation. Knockdown of lncRNA 4930581F22Rik hindered skeletal muscle differentiation and resulted in the inhibition of the myogenic markers MyHC and MEF2C. Furthermore, we found that lncRNA 4930581F22Rik regulates myogenesis via the ERK/MAPK signaling pathway, and this effect could be attenuated by the ERK-specific inhibitor PD0325901. Additionally, in vivo mice injury model results revealed that lncRNA 4930581F22Rik is involved in skeletal muscle regeneration. These results establish a theoretical basis for understanding the contribution of lncRNAs in skeletal muscle development and regeneration.

The fourth species of Leptobrachella (Anura, Megophryidae) found at Shiwandashan National Nature Reserve, Guangxi, China.

A new species of the genus Leptobrachella, L.guinanensissp. nov., is described in this study based on morphological, molecular, and bioacoustic data. The species was discovered in the Shiwandashan National Nature Reserve in Shangsi County, Guangxi, China. Phylogenetically, L.guinanensissp. nov. is closely related to L.ventripunctata. However, there are distinct morphological differences between L.guinanensissp. nov. and L.ventripunctata, as well as three other sympatric species (L.shangsiensis, L.shiwandashanensis, and L.sungi). These differences include body size (SVL 30.5-32.5 mm in males; 38.7-41.8 mm in females in the new species vs 25.5-28.0 mm in males, 31.5-35.0 mm in females in L.ventripunctata), the absence of brown spots on the ventral surface (vs chest and belly creamy white with many scattered brown spots in L.ventripunctata), 1/3 toe webbing and wide toe lateral fringes (vs no toe webbing and no lateral fringes in L.ventripunctata), and distinct dermal ridges under toes (vs absent in L.ventripunctata). Furthermore, the dominant vocal frequencies of the new species range from 7.3 to 8.3 kHz, which is unique compared to other Leptobrachella species and represents the highest dominant frequencies ever recorded. The Shiwandashan National Nature Reserve is now home to four known sympatric species of Leptobrachella.

[Molecular identification of animal species of Bufonis Venenum from secretions].

The animal species is one of the key factors affecting the quality of Bufonis Venenum. The quality of Bufonis Venenum derived from Bufo bufo gargarizans is significantly higher than that from B. melanostictus. Since Bufonis Venenum is from secretions, the conventional identification methods are difficult to identify the animal species due to the lack of the appearance and morphology of the animals. The rapid development of molecular identification technology has provided new methods for the identification of Bufonis Venenum. However, because of the low content and serve degradation of residual DNA in secretions, the research on the molecular identification of Chinese medicinal materials from secretions remains to be carried out. To understand the animal species of Bufonis Venenum, this study collected 83 samples of Bufonis Venenum, including 7 commercially available samples, 5 reference medicinal materials, and 71 animal samples from which Bufonis Venenum was prepared according to the method in the 2020 edition of the Chinese Pharmacopoeia. Different DNA extraction methods were used and compared, and the mitochondrial 16S rRNA gene fragments were amplified, on the basis of which the phylogenetic trees were built. Finally, molecular identification of the animal species of the samples was performed. The results showed that the DNA extracted from Bufonis Venenum by the reagent kit had good quality, and 16S rRNA sequences were successfully amplified from 80 out of the 83 samples. In addition, 71 16S rRNA sequences of the animal species of Bufonis Venenum were downloaded from GenBank. The phylogenetic trees constructed based on the neighbor-joining(NJ) method and the Bayesian inference(BI) method showed that the samples derived from B. bufo gargarizans and B. melanostictus were clustered into separate monophyletic clades, with the support of 100%(NJ) and 1.00(BI), respectively. The animal species of both commercially available samples and reference medicinal materials were B. bufo gargarizans. In conclusion, DNA can be extracted from Bufonis Venenum derived from secretions, and the 16S rRNA gene sequences can be amplified, which can be used for molecular identification of the animal species of Bufonis Venenum. The findings provide a reference for the quality control of Bufonis Venenum and the identification of animal species of medicinal materials derived from secretions.

A new species of Odorrana Fei, Ye & Huang, 1990 (Amphibia, Anura, Ranidae) from central Guangxi, China with a discussion of the taxonomy of Odorrana (Bamburana).

A new species of odorous frog, Odorranadamingshanensissp. nov., was found at the Damingshan National Nature Reserve in Guangxi, China. This species can be distinguished from its congeners by a combination of the following characters: medium body size (SVL 52.3-54.8 mm in males and 74.8-81.2 mm in females), sawtooth spinules on the upper lip, obtusely rounded snout that extends beyond the lower margin, distinct dorsolateral folds, horny tubercles on the rear of the back, presence of outer metatarsal tubercles, dilated nuptial pad with velvety spinules, distinct maxillary gland with tiny spines, and external lateral vocal sac. Through analysis of the 16S mitochondria gene, the new species is closely related to O.nasica and O.yentuensis, but the genetic divergence between the new species and the latter exceeds 7% (uncorrected p-distance). Currently, the new species is only known from its original discovery site. Furthermore, a discussion on the taxonomy of Odorrana (Bamburana) was conducted, identifying seven species within the subgenus Odorrana (Bamburana).

A new species of the genus Leptobrachella Smith 1925 (Amphibia, Anura, Megophryidae) from Guangxi, China.

A new species of Leptobrachella, L.wumingensissp. nov., was described from the Damingshan National Nature Reserve, Wuming District, Nanning City, Guangxi, China based on morphological, molecular and bioacoustic data. Phylogenetic analysis of 16S mtDNA fragments revealed that the new species is closely related to L.damingshanensis. Uncorrected p-distances between the new species and all homologous DNA sequences available for the 16S gene of Leptobrachella are greater than 7.1%. Morphologically, L.wumingensissp. nov. differs from its congeners in several ways, including a medium body size (SVL 26.0-26.7 mm in males, 30.6-34.8 mm in females), lack of toe webbing and lateral fringes, shagreened and granular dorsal surface, pale brown dorsum with darker brown markings, iris bicolored, with the upper half copper and fading to silver in the lower half, and the presence of small irregular black spots and tangerine tubercles on the flanks. Furthermore, we found the new species to have two types of advertisement calls and relatively high dominant frequencies, making it distinct from its congeners.

Live-cell imaging reveals redox metabolic reprogramming during zygotic genome activation.

Metabolic programming is deeply intertwined with early embryonic development including zygotic genome activation (ZGA), the polarization of zygotic cells, and cell fate commitment. It is crucial to establish a noninvasive imaging technology that spatiotemporally illuminates the cellular metabolism pathways in embryos to track developmental metabolism in situ. In this study, we used two high-quality genetically encoded fluorescent biosensors, SoNar for NADH/NAD+ and iNap1 for NADPH, to characterize the dynamic regulation of energy metabolism and redox homeostasis during early zygotic cleavage. Our imaging results showed that NADH/NAD+ levels decreased from the early to the late two-cell stage, whereas the levels of the reducing equivalent NADPH increased. Mechanistically, transcriptome profiling suggested that during the two-cell stage, zygotic cells downregulated the expression of genes involved in glucose uptake and glycolysis, and upregulated the expression of genes for pyruvate metabolism in mitochondria and oxidative phosphorylation, with a decline in the expression of two peroxiredoxin genes, Prdx1 and Prdx2. Collectively, with the establishment of in situ metabolic monitoring technology, our study revealed the programming of redox metabolism during ZGA.

ULK1-mediated metabolic reprogramming regulates Vps34 lipid kinase activity by its lactylation.

Autophagy and glycolysis are highly conserved biological processes involved in both physiological and pathological cellular programs, but the interplay between these processes is poorly understood. Here, we show that the glycolytic enzyme lactate dehydrogenase A (LDHA) is activated upon UNC-51-like kinase 1 (ULK1) activation under nutrient deprivation. Specifically, ULK1 directly interacts with LDHA, phosphorylates serine-196 when nutrients are scarce and promotes lactate production. Lactate connects autophagy and glycolysis through Vps34 lactylation (at lysine-356 and lysine-781), which is mediated by the acyltransferase KAT5/TIP60. Vps34 lactylation enhances the association of Vps34 with Beclin1, Atg14L, and UVRAG, and then increases Vps34 lipid kinase activity. Vps34 lactylation promotes autophagic flux and endolysosomal trafficking. Vps34 lactylation in skeletal muscle during intense exercise maintains muscle cell homeostasis and correlates with cancer progress by inducing cell autophagy. Together, our findings describe autophagy regulation mechanism and then integrate cell autophagy and glycolysis.

Redox metabolism maintains the leukemogenic capacity and drug resistance of AML cells.

Rewiring of redox metabolism has a profound impact on tumor development, but how the cellular heterogeneity of redox balance affects leukemogenesis remains unknown. To precisely characterize the dynamic change in redox metabolism in vivo, we developed a bright genetically encoded biosensor for H2O2 (named HyPerion) and tracked the redox state of leukemic cells in situ in a transgenic sensor mouse. A H2O2-low (HyPerion-low) subset of acute myeloid leukemia (AML) cells was enriched with leukemia-initiating cells, which were endowed with high colony-forming ability, potent drug resistance, endosteal rather than vascular localization, and short survival. Significantly high expression of malic enzymes, including ME1/3, accounted for nicotinamide adenine dinucleotide phosphate (NADPH) production and the subsequent low abundance of H2O2. Deletion of malic enzymes decreased the population size of leukemia-initiating cells and impaired their leukemogenic capacity and drug resistance. In summary, by establishing an in vivo redox monitoring tool at single-cell resolution, this work reveals a critical role of redox metabolism in leukemogenesis and a potential therapeutic target.

Ultrasensitive sensors reveal the spatiotemporal landscape of lactate metabolism in physiology and disease.

Despite its central importance in cellular metabolism, many details remain to be determined regarding subcellular lactate metabolism and its regulation in physiology and disease, as there is sensitive spatiotemporal resolution of lactate distribution, and dynamics remains a technical challenge. Here, we develop and characterize an ultrasensitive, highly responsive, ratiometric lactate sensor, named FiLa, enabling the monitoring of subtle lactate fluctuations in living cells and animals. Utilizing FiLa, we demonstrate that lactate is highly enriched in mammalian mitochondria and compile an atlas of subcellular lactate metabolism that reveals lactate as a key hub sensing various metabolic activities. In addition, FiLa sensors also enable direct imaging of elevated lactate levels in diabetic mice and facilitate the establishment of a simple, rapid, and sensitive lactate assay for point-of-care clinical screening. Thus, FiLa sensors provide powerful, broadly applicable tools for defining the spatiotemporal landscape of lactate metabolism in health and disease.

Role of lncRNA Has2os in Skeletal Muscle Differentiation and Regeneration.

Long non-coding RNAs (lncRNAs) regulate a series of physiological processes and play an important role in development, metabolism and disease. Our previous studies showed that lncRNAs involved in skeletal muscle differentiation. Here, we demonstrated that lncRNA Has2os is highly expressed in skeletal muscle and significantly elevated during skeletal cell differentiation. The knockdown of Has2os inhibited myocyte fusion and impeded the expression of the myogenic factors MyHC and Mef2C. Mechanically, Has2os regulates skeletal muscle differentiation by inhibiting the JNK/MAPK signaling pathway. Furthermore, we also revealed that Has2os is involved in the early stage of regeneration after muscle injury, and the JNK/MAPK signaling pathway is activated at both protein and mRNA levels during early repair. Our results demonstrate the new function of lncRNA Has2os, which plays crucial roles during skeletal muscle differentiation and muscle regeneration, providing a basis for the therapy of lncRNA-related muscle diseases.

Phylogenetic analysis of combined mitochondrial genome and 32 nuclear genes provides key insights into molecular systematics and historical biogeography of Asian warty newts of the genus Paramesotriton (Caudata: Salamandridae).

The Paramesotriton Chang, 1935 genus of Asian warty newts is the second most diverse genus in the family Salamandridae, currently containing 14 recognized species from northern Vietnam to southwest-central and southern China. Although species of this genus have been included in previous phylogenetic studies, the origin and interspecific relationships of the genus are still not fully resolved, especially at key nodes in the phylogeny. In this study, we sequenced mitochondrial genomes and 32 nuclear genes from 27 samples belonging to 14 species to reconstruct the interspecific phylogenetic relationships within Paramesotriton and explore its historical biogeography in southern China. Both Bayesian inference and maximum-likelihood analyses highly supported the monophyly of Paramesotriton and its two recognized species groups ( P. caudopunctatus and P. chinensis groups) and further identified five hypothetical phylogenetic cryptic species. Biogeographic analyses indicated that Paramesotriton originated in southwestern China (Yunnan-Guizhou Plateau/South China) during the late Oligocene. The time of origin of Paramesotriton corresponded to the second uplift of the Himalayan/Qinghai-Xizang (Tibetan) Plateau (QTP), rapid lateral extrusion of Indochina, and formation of karst landscapes in southwestern China. Principal component analysis (PCA), independent sample t-tests, and niche differentiation using bioclimatic variables based on locations of occurrence suggested that Paramesotriton habitat conditions in the three current regions (West, South, and East) differ significantly, with different levels of climatic niche differentiation. Species distribution model (SDM) predictions indicated that the most suitable distribution areas for the P. caudopunctatus and P. chinensis species groups are western and southern/eastern areas of southern China. This study increases our knowledge of the taxonomy, biodiversity, origin, and suitable distribution areas of the genus Paramesotriton based on phylogenetic, biogeographic, and species distribution models.

NADPH metabolism determines the leukemogenic capacity and drug resistance of AML cells.

The mechanism by which redox metabolism regulates the fates of acute myeloid leukemia (AML) cells remains largely unknown. Using a highly sensitive, genetically encoded fluorescent sensor of nicotinamide adenine dinucleotide phosphate (NADPH), iNap1, we find three heterogeneous subpopulations of AML cells with different cytosolic NADPH levels in an MLL-AF9-induced murine AML model. The iNap1-high AML cells have enhanced proliferation capacities both in vitro and in vivo and are enriched for more functional leukemia-initiating cells than iNap1-low counterparts. The iNap1-high AML cells prefer localizing in the bone marrow endosteal niche and are resistant to methotrexate treatment. Furthermore, iNap1-high human primary AML cells have enhanced proliferation abilities both in vitro and in vivo. Mechanistically, the MTHFD1-mediated folate cycle regulates NADPH homeostasis to promote leukemogenesis and methotrexate resistance. These results provide important clues for understanding mechanisms by which redox metabolism regulates cancer cell fates and a potential metabolic target for AML treatments.

Spatiotemporal monitoring of NAD+ metabolism with fluorescent biosensors.

Nicotinamide adenine dinucleotide (NAD+) plays vital roles in cell metabolism, cell signaling, and gene expression regulation. NAD+ shows intricate subcellular distribution and dynamic changes in diverse biological processes; however, traditional biochemical assays usually require cell lysis, making it technically difficult to measure NAD+ metabolism in live cells or in vivo. Recently, a few genetically encoded and semisynthetic fluorescent sensors have been used to monitor NAD+ metabolism in a variety of biological settings. In this review, we summarize the recent progress in the development of fluorescent sensors for NAD+ and their applications in life science research. We further analyze their advantages, limitations, and perspectives for future development.

Identification of a seven-gene prognostic signature using the gene expression profile of osteosarcoma.

Background: Osteosarcoma is a malignant bone tumor that typically occurs in adolescents or children under 20 years of age. Developing efficient clinical prognostic markers is crucial for improving the treatment of osteosarcoma patients. Methods: Three datasets related to osteosarcoma were acquired from the Gene Expression Omnibus (GEO) database. A gene signature model was established using the Limma package in the R software, univariate and multivariate survival analyses, and least absolute shrinkage and selection operator (LASSO) algorithms. The gene signature was then verified using external datasets. Results: From the GEO database, 242 differentially expressed genes were identified. A total of 590 unique genes, including 380 genes from the human protein interaction network, were found to be related to biological processes such as bone development and bone cell development. Univariate Cox survival analyses revealed 43 genes that were associated with the prognosis of osteosarcoma patients. A seven-gene signature [retinitis pigmentosa 2 (RP2), polyhydroxybutyrate (PHB), myosin VI (MYO6), mutL homolog 1 (MLH1), Casein kinase 2 beta (CSNK2B), ribosomal protein L37A (RPL37A), and CCAAT/enhancer binding protein alpha (CEBPA)] was developed using LASSO regression analysis and multivariate regression analysis. This gene signature could stratify the prognostic risk of sample cases in the training set, the test set, and the external verification set (P<0.01). The area under the receiver operating characteristic curve for the 5-year survival was higher than 0.72 in both the training and verification groups. Conclusions: In this study, a seven-gene signature was developed that is highly efficient at predicting the prognosis of patients with osteosarcoma, and therefore, this signature may be a crucial guide in the treatment of these patients.

A new species of Nidirana (Anura, Ranidae) from northern Guangxi, China.

A new species of music frog, Nidiranaguibeiensis sp. nov., is described from northern Guangxi, China. Based on two mtDNA fragments analyzed, phylogenetic trees reveal that N.guibeiensis sp. nov. is most closely related to N.leishanensis. However, the new species can be identified by conspicuous diagnostic morphological characteristics as well as bioacoustics. In contrast to the known Nidirana species, the advertisement calls of the new species can be divided into three types, calls with one, two, and three notes. In addition, the new species has nest construction behavior, which is inconsistent with N.leishanensis. Nidiranaguibeiensis sp. nov. occurs in paddy fields or still pools at 300-1300 m a.s.l.

A new species of the genus Rana sensu lato Linnaeus, 1758 (Anura, Ranidae) from Wuyi Mountain, Fujian Province, China.

A new species of the frog genus Rana sensu lato from Wuyi Mountain, Fujian Province, China is described. Molecular phylogenetic analyses clustered the new species into the R.johnsi group and indicated that it was genetically divergent from its closely related species. The new species could be distinguished from its congeners by a combination of the following characters: body size medium, SVL 41.4-45.6 mm (42.9 ± 1.9 mm, n = 4) in adult males and 47.6-50.3 mm (n = 2) in adult females; adult male with a pair of internal subgular vocal sacs; lateroventral grooves present on tip of toes; webbing on fourth toes reaching the tip of toe; transverse skin ridges distinctly present on the dorsal surface of thigh and tibia, the number large (mean 26.5 ± 2.7, range 22-29, n = 6); breeding males possess creamy white nuptial pad with tiny velvety spines on the dorsal surface of the first finger, divided into three parts.

A new species of Leptobrachella Smith 1925 (Anura: Megophryidae) from Southern Guangxi, China.

We described a new species of the genus Leptobrachella from Southern Guangxi, China, based on morphological characteristics, molecular analyses and bioacoustics. This new species, Leptobrachella shiwandashanensis sp. nov., occurs in sympatry with Leptobrachella shangsiensis. However, phylogenetic analyses based on the mitochondrial 16S gene fragment revealed that the new species was clustered in a monophyletic group and a considerable genetic divergence existed between the new species and L. shangsiensis (p-distance: 7.9%) as well as its congeners (minimum p-distance: 7.0%). The new species also differed from its congeners by a combination of the following characteristics: (1) small size (SVL 26.829.7 mm in males; 33.735.9 mm in females); (2) pale brown dorsal surfaces, with a brown inverse-triangle-shaped marking between the eyes; (3) creamy white ventral surface with brown spots on the lateral margin, with a near immaculate creamy white throat and chest; (4) without webbing and lateral fringes on toes; (5) flanks with irregular black spots; (6) tibia-tarsal articulation reaching the posterior of the eye in males but reaching the shoulder in females; (7) heels that do not meet when the thighs are appressed at right angles to the body; (8) bicolored iris, with the upper half brownish-red, fading to silver in the lower half; and (9) a call consisting of a single note and a dominant frequency of 5.35.7 kHz (recorded at 23C).

Characterization of Long Non-coding RNAs Modified by m6A RNA Methylation in Skeletal Myogenesis.

Proper development of mammalian skeletal muscle relies on precise gene expression regulation. Our previous studies revealed that muscle development is regulated by both mRNA and long non-coding RNAs (lncRNAs). Accumulating evidence has demonstrated that N6-methyladenosine (m6A) plays important roles in various biological processes, making it essential to profile m6A modification on a transcriptome-wide scale in developing muscle. Patterns of m6A methylation in lncRNAs in developing muscle have not been uncovered. Here, we reveal differentially expressed lncRNAs and report temporal m6A methylation patterns in lncRNAs expressed in mouse myoblasts and myotubes by RNA-seq and methylated RNA immunoprecipitation (MeRIP) sequencing. Many lncRNAs exhibit temporal differential expression, and m6A-lncRNAs harbor the consensus m6A motif "DRACH" along lncRNA transcripts. Interestingly, we found that m6A methylation levels of lncRNAs are positively correlated with the transcript abundance of lncRNAs. Overexpression or knockdown of m6A methyltransferase METTL3 alters the expression levels of these lncRNAs. Furthermore, we highlight that the function of m6A genic lncRNAs might correlate to their nearby mRNAs. Our work reveals a fundamental expression reference of m6A-mediated epitranscriptomic modifications in lncRNAs that are temporally expressed in developing muscle.