Adipocyte subpopulations mediate lipolysis and obesity-induced insulin resistance.

Studies in humans and mice have determined that distinct subpopulations of adipocytes reside even within individual adipose tissue depots. Previously, our lab defined three white adipocyte subpopulations with stable and unique gene expression profiles, which were termed type 1, 2, and 3 adipocytes, respectively. Our previous studies demonstrated that type 2 adipocytes were highly responsive to the inflammatory cytokine, tumor necrosis factor alpha (TNFα). This study extends these findings to investigate the role of type 2 adipocytes in obesity. We found that treatment with TNFα increased lipolysis specifically in type 2 adipocytes, at least in part, through the reduction of fat-specific protein 27 (FSP27) expression. To assess the physiological role of lipolysis from this adipocyte subpopulation, a type2Ad-hFSP27tg mouse model was generated by overexpressing human FSP27 specifically in type 2 adipocytes. Glucose and insulin tolerance test analysis showed that male type2Ad-hFSP27tg mice on 60% high-fat diet exhibited improved glucose tolerance and insulin sensitivity, with no change in body weight compared to controls. These metabolic changes may, at least in part, be explained by the reduced lipolysis rate in the visceral fat of type2Ad-hFSP27tg mice. Although FSP27 overexpression in primary type 2 adipocytes was sufficient to acutely reduce TNFα-induced apoptosis in vitro, it failed to reduce macrophage infiltration in obesity in vivo. Taken together, these results strongly suggest that type 2 adipocytes contribute to the regulation of lipolysis and could serve as a potential therapeutic target for obesity-associated insulin resistance.

Adipocyte Subpopulations Mediate Growth Hormone-induced Lipolysis and Glucose Tolerance in Male Mice.

In adipose tissue, growth hormone (GH) stimulates lipolysis, leading to an increase in plasma free fatty acid levels and a reduction in insulin sensitivity. In our previous studies, we have found that GH increases lipolysis by reducing peroxisome proliferator-activated receptor γ (PPARγ) transcription activity, leading to a reduction of tat-specific protein 27 (FSP27, also known as CIDEC) expression. In previous studies, our laboratory uncovered 3 developmentally distinct subpopulations of white adipocytes. In this manuscript, we show that one of the subpopulations, termed type 2 adipocytes, has increased GH-induced signaling and lipolysis compared to other adipocyte subtypes. To assess the physiological role of GH-mediated lipolysis mediated by this adipocyte subpopulation, we specifically expressed human FSP27 (hFSP27) transgene in type 2 adipocytes (type2Ad-hFSP27tg mice). Systemically, male type2Ad-hFSP27tg mice displayed reduced serum glycerol release and nonesterified fatty acids levels after acute GH treatment, and improvement in acute, but not chronic, GH-induced glucose intolerance. Furthermore, we demonstrate that type2Ad-hFSP27tg mice displayed improved hepatic insulin signaling. Taken together, these results indicate that this adipocyte subpopulation is a critical regulator of the GH-mediated lipolytic and metabolic response. Thus, further investigation of adipocyte subpopulations may provide novel treatment strategies to regulate GH-induced glucose intolerance in patients with growth and metabolic disorders.

Data-driven approaches to improve water-use efficiency and drought resistance in crop plants.

With the increasing population, there lies a pressing demand for food, feed and fibre, while the changing climatic conditions pose severe challenges for agricultural production worldwide. Water is the lifeline for crop production; thus, enhancing crop water-use efficiency (WUE) and improving drought resistance in crop varieties are crucial for overcoming these challenges. Genetically-driven improvements in yield, WUE and drought tolerance traits can buffer the worst effects of climate change on crop production in dry areas. While traditional crop breeding approaches have delivered impressive results in increasing yield, the methods remain time-consuming and are often limited by the existing allelic variation present in the germplasm. Significant advances in breeding and high-throughput omics technologies in parallel with smart agriculture practices have created avenues to dramatically speed up the process of trait improvement by leveraging the vast volumes of genomic and phenotypic data. For example, individual genome and pan-genome assemblies, along with transcriptomic, metabolomic and proteomic data from germplasm collections, characterised at phenotypic levels, could be utilised to identify marker-trait associations and superior haplotypes for crop genetic improvement. In addition, these omics approaches enable the identification of genes involved in pathways leading to the expression of a trait, thereby providing an understanding of the genetic, physiological and biochemical basis of trait variation. These data-driven gene discoveries and validation approaches are essential for crop improvement pipelines, including genomic breeding, speed breeding and gene editing. Herein, we provide an overview of prospects presented using big data-driven approaches (including artificial intelligence and machine learning) to harness new genetic gains for breeding programs and develop drought-tolerant crop varieties with favourable WUE and high-yield potential traits.

Emerging roles of long non-coding RNAs in regulating agriculturally important seed traits.

Seeds have enormous economic importance as source of calories, nutrition, edible oil, and biofuels. Therefore, seed traits like seed size and shape, weight, micronutrient content, oil content, quality, post-harvest sprouting, etc., are some of the main targets in crop improvement. Designing the strategies for their improvement benefits heavily from understanding the regulatory aspects of seed development. Recent studies indicate that long non-coding RNAs (lncRNAs) are one of the important regulators of seed development. They played a significant role in crop domestication by influencing seed traits. LncRNAs are conventionally defined as non-coding RNAs greater than 200 bp in length but lacking protein coding potential. Here we highlight the emerging pieces of evidence of lncRNA-mediated regulation of seed development through diverse mechanisms, for instance, by acting as target mimics or precursors of regulatory small RNAs or through chromatin remodeling and post-transcriptional repression. We also enumerate the insights from high-throughput transcriptomic studies from developing seeds of cereal, oilseed, biofuel, and pulse crops. We highlight the lncRNA candidates and lncRNA-mediated regulatory networks regulating seed development and related agronomic traits. Further, we discuss the potential of lncRNAs for improvement of agriculturally important seed traits through marker-assisted breeding and/or transgenic approaches.

Non-coding RNAs-mediated environmental surveillance determines male fertility in plants.

Plants are continuously exposed to environmental stresses leading to significant yield losses. With the changing climatic conditions, the intensity and duration of these stresses are expected to increase, posing a severe threat to crop productivity worldwide. Male gametogenesis is one of the most sensitive developmental stages. Exposure to environmental stresses during this stage leads to male sterility and yield loss. Elucidating the underlying molecular mechanism of environment-affected male sterility is essential to address this challenge. High-throughput RNA sequencing studies, loss-of-function phenotypes of sRNA biogenesis genes and functional genomics studies with non-coding RNAs have started to unveil the roles of small RNAs, long non-coding RNAs and the complex regulatory interactions between them in regulating male fertility under different growth regimes. Here, we discuss the current understanding of the non-coding RNA-mediated environmental stress surveillance and regulation of male fertility in plants. The candidate ncRNAs emerging from these studies can be leveraged to generate environment-sensitive male sterile lines for hybrid breeding or mitigate the impact of climate change on male fertility, as the situation demands.

Comment on Muth et al. Assessing Critical Flicker Fusion Frequency: Which Confounders? A Narrative Review. Medicina 2023, 59, 800.

We first want to thank the authors of the excellent review for their contributions to summarizing the confounders associated with critical flicker fusion frequency (CFFF) [...].

Cognitive Functions in Scuba, Technical and Saturation Diving.

Scuba diving as a recreational activity is becoming increasingly popular. However, the safety of this activity, especially in the out-of-comfort zone, has been discussed worldwide. The latest publications bring conclusions regarding negative effects on cognitive functions. We compare the acute and chronic effects of diving on cognitive functioning depending on the type of dive performed, including recreational, technical and saturation diving. However, the results of research show that acute and chronic effects on cognitive functions can be negative. While acute effects are reversible after the ascent, chronic effects include white matter lesions in magnetic resonance imaging scans. We believe that more investigations should be performed to determine the chronic effects that could be observed after a few months of observations in a group of regular, intense divers. In addition, publications referring to technical divers are very limited, which is disquieting, as this particular group of divers seems to be neglected in research concerning the effects of diving on cognitive functions.

Transcriptome analysis reveals cell cycle-related transcripts as key determinants of varietal differences in seed size of Brassica juncea.

Brassica juncea is an important oilseed crop, widely grown as a source of edible oil. Seed size is a pivotal agricultural trait in oilseed Brassicas. However, the regulatory mechanisms underlying seed size determination are poorly understood. To elucidate the transcriptional dynamics involved in the determination of seed size in B. juncea, we performed a comparative transcriptomic analysis using developing seeds of two varieties, small-seeded Early Heera2 (EH2) and bold-seeded Pusajaikisan (PJK), at three distinct stages (15, 30 and 45 days after pollination). We detected 112,550 transcripts, of which 27,186 and 19,522 were differentially expressed in the intra-variety comparisons and inter-variety comparisons, respectively. Functional analysis using pathway, gene ontology, and transcription factor enrichment revealed that cell cycle- and cell division-related transcripts stay upregulated during later stages of seed development in the bold-seeded variety but are downregulated at the same stage in the small-seeded variety, indicating that an extended period of cell proliferation in the later stages increased seed weight in PJK as compared to EH2. Further, k-means clustering and candidate genes-based analyses unravelled candidates for employing in seed size improvement of B. juncea. In addition, candidates involved in determining seed coat color, oil content, and other seed traits were also identified.

Reply to Vrijdag et al. Comment on "Mankowska et al. Critical Flicker Fusion Frequency: A Narrative Review. Medicina 2021, 57, 1096".

Thank you very much for your interest and comments [...].

Spatiotemporal Regulation of CRISPR/Cas9 Enables Efficient, Precise, and Heritable Edits in Plant Genomes.

CRISPR/Cas-mediated editing has revolutionized crop engineering. Due to the broad scope and potential of this technology, many studies have been carried out in the past decade towards optimizing genome editing constructs. Clearly, the choice of the promoter used to drive gRNA and Cas9 expression is critical to achieving high editing efficiency, precision, and heritability. While some important considerations for choosing a promoter include the number and nature of targets, host organism, mode of transformation and goal of the experiment, spatiotemporal regulation of Cas9 expression using tissue-specific or inducible promoters enables higher heritability and efficiency of targeted mutagenesis with reduced off-target effects. In this review, we discuss specific studies that highlight the prospects and trade-offs associated with the choice of promoters on genome editing and emphasize the need for inductive exploration and discovery to further advance this area of research in crop plants.

Insights into the molecular basis of hypergravity-induced root growth phenotype in bread wheat (Triticum aestivum L.).

Hypergravity is a condition where the force of gravity exceeds that on the surface of the Earth and can be simulated by centrifugation. Previously, a significant increase in root growth phenotype was observed when wheat seeds were exposed to hypergravity (10 g for 12 h). In the present study, we investigated the molecular basis of this change through root transcriptome. The data revealed a total of 3765 up-regulated and 2102 down-regulated transcripts in response to hypergravity. GO enrichment analysis revealed hormonal responses, cell division, and cell-wall-related terms were significantly enriched in hypergravity. The increased isoform level expression of transcripts involved in auxin biosynthesis, transport, and signaling was observed. Further, enhanced expression of cell division transcripts and down-regulation of cell number regulator genes suggests rapid cell division. Overexpression of cellulose and hemicellulose biosynthesis transcripts suggests demand for cell-wall constituents. Collectively, this study identified candidate genes associated with hypergravity-induced enhanced root growth.

Genome-wide in silico analysis of long intergenic non-coding RNAs from rice peduncles at the heading stage.

Long intergenic non-coding RNAs (lincRNAs) belong to the category of long non-coding RNAs (lncRNAs), originated from intergenic regions, which do not code for proteins. LincRNAs perform prominent role in regulation of gene expression during plant development and stress response by directly interacting with DNA, RNA, or proteins, or triggering production of small RNA regulatory molecules. Here, we identified 2973 lincRNAs and investigated their expression dynamics during peduncle elongation in two Indian rice cultivars, Pokkali and Swarna, at the time of heading. Differential expression analysis revealed common and cultivar-specific expression patterns, which we utilized to infer the lincRNA candidates with potential involvement in peduncle elongation and panicle exsertion. Their putative targets were identified using in silico prediction methods followed by pathway mapping and literature-survey based functional analysis. Further, to infer the mechanism of action, we identified the lincRNAs which potentially act as miRNA precursors or target mimics. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01059-2.

Critical Flicker Fusion Frequency: A Narrative Review.

This review presents the current knowledge of the usage of critical flicker fusion frequency (CFF) in human and animal model studies. CFF has a wide application in different fields, especially as an indicator of cortical arousal and visual processing. In medicine, CFF may be helpful for diagnostic purposes, for example in epilepsy or minimal hepatic encephalopathy. Given the environmental studies and a limited number of other methods, it is applicable in diving and hyperbaric medicine. Current research also shows the relationship between CFF and other electrophysiological methods, such as electroencephalography. The human eye can detect flicker at 50-90 Hz but reports are showing the possibility to distinguish between steady and modulated light up to 500 Hz. Future research with the use of CFF is needed to better understand its utility and application.

Divergence among rice cultivars reveals roles for transposition and epimutation in ongoing evolution of genomic imprinting.

Parent-of-origin-dependent gene expression in mammals and flowering plants results from differing chromatin imprints (genomic imprinting) between maternally and paternally inherited alleles. Imprinted gene expression in the endosperm of seeds is associated with localized hypomethylation of maternally but not paternally inherited DNA, with certain small RNAs also displaying parent-of-origin-specific expression. To understand the evolution of imprinting mechanisms in Oryza sativa (rice), we analyzed imprinting divergence among four cultivars that span both japonica and indica subspecies: Nipponbare, Kitaake, 93-11, and IR64. Most imprinted genes are imprinted across cultivars and enriched for functions in chromatin and transcriptional regulation, development, and signaling. However, 4 to 11% of imprinted genes display divergent imprinting. Analyses of DNA methylation and small RNAs revealed that endosperm-specific 24-nt small RNA-producing loci show weak RNA-directed DNA methylation, frequently overlap genes, and are imprinted four times more often than genes. However, imprinting divergence most often correlated with local DNA methylation epimutations (9 of 17 assessable loci), which were largely stable within subspecies. Small insertion/deletion events and transposable element insertions accompanied 4 of the 9 locally epimutated loci and associated with imprinting divergence at another 4 of the remaining 8 loci. Correlating epigenetic and genetic variation occurred at key regulatory regions-the promoter and transcription start site of maternally biased genes, and the promoter and gene body of paternally biased genes. Our results reinforce models for the role of maternal-specific DNA hypomethylation in imprinting of both maternally and paternally biased genes, and highlight the role of transposition and epimutation in rice imprinting evolution.

Development of an Optimized Clearing Protocol to Examine Adipocyte Subpopulations in White Adipose Tissue.

Organic solvent dibenzyl ether (DBE)-based protocols have been widely used in adipose tissue clearing. However, benzyl alcohol/benzyl benzoate (BABB)-based clearing has been shown to offer better transparency in other tissues. The addition of diphenyl ether (DPE) to BABB (BABB-D4) is often included to preserve fluorescent signals, but its effects on adipose tissue transparency and shrinkage have not been explored. Distinct adipocyte subpopulations contribute to its cellular composition and biological activity. Here, we compared clearing solvents to create an optimized clearing methodology for the study of adipocyte subpopulations. Adipose tissues were cleared with BABB, BABB-D4, and DBE, and post-clearing transparency and tissue shrinkage were measured. An optimized protocol, including BABB-D4 clearing, delipidation, and extensive immunofluorescence blocking steps, was created to examine the spatial distribution of Wt-1 positive progenitor-derived (Type-1) adipocytes in intact mesenteric fat. Both BABB and BABB-D4 lead to significantly increased tissue transparency with reduced tissue shrinkage compared to DBE-cleared adipose tissue. Type-1 adipocytes are found in a clustered distribution with predominant residence in fat associated with the ileum and colon. This paper details an optimized clearing methodology for adipose tissue with increased tissue transparency and reduced shrinkage, and therefore will be a useful tool for investigating adipose tissue biology.

MicroRNA-mediated regulation of agronomically important seed traits: a treasure trove with shades of grey!

Seed development is an intricate process with multiple levels of regulation. MicroRNAs (miRNAs) have emerged as one of the crucial components of molecular networks underlying agronomically important seed traits in diverse plant species. In fact, loss of function of the genes regulating miRNA biogenesis also exhibits defects in seed development. A total of 21 different miRNAs have experimentally been shown to regulate seed size, nutritional content, vigor, and shattering, and have been reviewed here. The mechanism details of the associated regulatory cascades mediated through transcriptional regulators, phytohormones, basic metabolic machinery, and secondary siRNAs are elaborated. Co-localization of miRNAs and their target regions with seed-related QTLs provides new avenues for engineering these traits using conventional breeding programs or biotechnological interventions. While global analysis of miRNAs using small RNA sequencing studies are expanding the repertoire of candidate miRNAs, recent revelations on their inheritance, transport, and mechanism of action would be instrumental in designing better strategies for optimizing agronomically relevant seed traits.

Genomic mechanisms of climate adaptation in polyploid bioenergy switchgrass.

Long-term climate change and periodic environmental extremes threaten food and fuel security1 and global crop productivity2-4. Although molecular and adaptive breeding strategies can buffer the effects of climatic stress and improve crop resilience5, these approaches require sufficient knowledge of the genes that underlie productivity and adaptation6-knowledge that has been limited to a small number of well-studied model systems. Here we present the assembly and annotation of the large and complex genome of the polyploid bioenergy crop switchgrass (Panicum virgatum). Analysis of biomass and survival among 732 resequenced genotypes, which were grown across 10 common gardens that span 1,800 km of latitude, jointly revealed extensive genomic evidence of climate adaptation. Climate-gene-biomass associations were abundant but varied considerably among deeply diverged gene pools. Furthermore, we found that gene flow accelerated climate adaptation during the postglacial colonization of northern habitats through introgression of alleles from a pre-adapted northern gene pool. The polyploid nature of switchgrass also enhanced adaptive potential through the fractionation of gene function, as there was an increased level of heritable genetic diversity on the nondominant subgenome. In addition to investigating patterns of climate adaptation, the genome resources and gene-trait associations developed here provide breeders with the necessary tools to increase switchgrass yield for the sustainable production of bioenergy.

Gene Expression Dynamics in Rice Peduncles at the Heading Stage.

Improving grain yield in the staple food crop rice has been long sought goal of plant biotechnology. One of the traits with significant impact on rice breeding programs is peduncle elongation at the time of heading failing which leads to significant reduction in grain yield due to incomplete panicle exsertion. To decipher transcriptional dynamics and molecular players underlying peduncle elongation, we performed RNA sequencing analysis of elongating and non-elongating peduncles in two Indian cultivars, Swarna and Pokkali, at the time of heading. Along with genes associated with cell division and cell wall biosynthesis, we observed significant enrichment of genes associated with auxins, gibberellins, and brassinosteroid biosynthesis/signaling in the elongating peduncles before heading in both the genotypes. Similarly, genes associated with carbohydrate metabolism and mobilization, abiotic stress response along with cytokinin, abscisic acid, jasmonic acid, and ethylene biosynthesis/signaling were enriched in non-elongating peduncles post heading. Significant enrichment of genes belonging to key transcription factor families highlights their specialized roles in peduncle elongation and grain filling before and after heading, respectively. A comparison with anther/pollen development-related genes provided 76 candidates with overlapping roles in anther/pollen development and peduncle elongation. Some of these are important for carbohydrate remobilization to the developing grains. These can be engineered to combat with incomplete panicle exsertion in male sterile lines and manipulate carbohydrate dynamics in grasses. Overall, this study provides baseline information about potential target genes for engineering peduncle elongation with implications on plant height, biomass composition and grain yields in rice.