Organ-specific Expression Patterns Research Articles

The mouse multi-organ proteome from infancy to adulthood

The early-life organ development and maturation shape the fundamental blueprint for later-life phenotype. However, a multi-organ proteome atlas from infancy to adulthood is currently not available. Herein, we present a comprehensive proteomic analysis of ten mouse organs (brain, heart, lung, liver, kidney, spleen, stomach, intestine, muscle and skin) at three crucial developmental stages (1-, 4- and 8-weeks after birth) acquired using data-independent acquisition mass spectrometry. We detect and quantify 11,533 protein groups across the ten organs and obtain 115 age-related differentially expressed protein groups that are co-expressed in all organs from infancy to adulthood. We find that spliceosome proteins prevalently play crucial regulatory roles in the early-life development of multiple organs, and detect organ-specific expression patterns and sexual dimorphism. This multi-organ proteome atlas provides a fundamental resource for understanding the molecular mechanisms underlying early-life organ development and maturation.

Genomic identification and expression analysis of acid invertase (AINV) gene family in Dendrobium officinale Kimura et Migo

BackgroundDendrobium officinale Kimura et Migo, a renowned traditional Chinese orchid herb esteemed for its significant horticultural and medicinal value, thrives in adverse habitats and contends with various abiotic or biotic stresses. Acid invertases (AINV) are widely considered enzymes involved in regulating sucrose metabolism and have been revealed to participate in plant responses to environmental stress. Although members of AINV gene family have been identified and characterized in multiple plant genomes, detailed information regarding this gene family and its expression patterns remains unknown in D. officinale, despite their significance in polysaccharide biosynthesis.ResultsThis study systematically analyzed the D. officinale genome and identified four DoAINV genes, which were classified into two subfamilies based on subcellular prediction and phylogenetic analysis. Comparison of gene structures and conserved motifs in DoAINV genes indicated a high-level conservation during their evolution history. The conserved amino acids and domains of DoAINV proteins were identified as pivotal for their functional roles. Additionally, cis-elements associated with responses to abiotic and biotic stress were found to be the most prevalent motif in all DoAINV genes, indicating their responsiveness to stress. Furthermore, bioinformatics analysis of transcriptome data, validated by quantitative real-time reverse transcription PCR (qRT-PCR), revealed distinct organ-specific expression patterns of DoAINV genes across various tissues and in response to abiotic stress. Examination of soluble sugar content and interaction networks provided insights into stress release and sucrose metabolism.ConclusionsDoAINV genes are implicated in various activities including growth and development, stress response, and polysaccharide biosynthesis. These findings provide valuable insights into the AINV gene amily of D. officinale and will aid in further elucidating the functions of DoAINV genes.

AtMYB41 acts as a dual-function transcription factor that regulates the formation of lipids in an organ- and development-dependent manner.

The plant cuticle controls non-stomatal water loss and can serve as a barrier against biotic agents, whereas the heteropolymer suberin and its associated waxes are deposited constitutively at specific cell wall locations. While several transcription factors controlling cuticle formation have been identified, those involved in the transcriptional regulation of suberin biosynthesis remain poorly characterized. The major goal of this study was to further analyse the function of the R2R3-Myeloblastosis (MYB) transcription factor AtMYB41 in formation of the cuticle, suberin, and suberin-associated waxes throughout plant development. For functional analysis, the organ-specific expression pattern of AtMYB41 was analysed and Atmyb41ge alleles were generated using the CRISPR/Cas9 system. These were investigated for root growth and water permeability upon stress. In addition, the fatty acid, wax, cutin, and suberin monomer composition of different organs was evaluated by gas chromatography. The characterization of Atmyb41ge mutants revealed that AtMYB41 negatively regulates the production of cuticular lipids and fatty acid biosynthesis in leaves and seeds, respectively. Remarkably, biochemical analyses indicate that AtMYB41 also positively regulates the formation of cuticular waxes in stems of Arabidopsis thaliana. Overall, these results suggest that the AtMYB41 acts as a negative regulator of cuticle and fatty acid biosynthesis in leaves and seeds, respectively, but also as a positive regulator of wax production in A. thaliana stems.

NnARF17 and NnARF18 from lotus promote root formation and modulate stress tolerance in transgenic Arabidopsis thaliana

Auxin response factors (ARFs) play a crucial role in regulating gene expression within the auxin signal transduction pathway, particularly during adventitious root (AR) formation. In this investigation, we identified full-length sequences for ARF17 and ARF18, encompassing 1,800 and 2,055 bp, encoding 599 and 684 amino acid residues, respectively. Despite exhibiting low sequence homology, the ARF17- and ARF18-encoded proteins displayed significant structural similarity and shared identical motifs. Phylogenetic analysis revealed close relationships between NnARF17 and VvARF17, as well as NnARF18 and BvARF18. Both ARF17 and ARF18 demonstrated responsiveness to exogenous indole-3-acetic acid (IAA), ethephon, and sucrose, exhibiting organ-specific expression patterns. Beyond their role in promoting root development, these ARFs enhanced stem growth and conferred drought tolerance while mitigating waterlogging stress in transgenic Arabidopsis plants. RNA sequencing data indicated upregulation of 51 and 75 genes in ARF17 and ARF18 transgenic plants, respectively, including five and three genes associated with hormone metabolism and responses. Further analysis of transgenic plants revealed a significant decrease in IAA content, accompanied by a marked increase in abscisic acid content under normal growth conditions. Additionally, lotus seedlings treated with IAA exhibited elevated levels of polyphenol oxidase, IAA oxidase, and peroxidase. The consistent modulation of IAA content in both lotus and transgenic plants highlights the pivotal role of IAA in AR formation in lotus seedlings.

Identification and organ-specific patterns of expression of two metallothioneins in the sentinel species Gammarus fossarum.

Metal pollution is a major concern for aquatic environments. Widespread contamination by various trace metal ions has been described in freshwater streams as well as their subsequent bioaccumulation, potentially leading to toxicity and trophic transfer. Metallothioneins constitute an evolutionary conserved family of low molecular weight, cysteine-rich, metal-chelating proteins, whose known physiological functions are the maintenance of the homeostasis of essential metals, the detoxification of non-essential metals, and the protection against oxidative stress and free radicals. In this study, we identified two metallothionein-coding transcripts, mt1 and mt2, in the transcriptome of the amphipod Gammarus fossarum, a sentinel species widely used to assess the quality of watersheds. For the first time, we investigated the organ-specific patterns of expression of these two mt transcripts at the individual level in the gills and the caeca of this small crustacean. In silico analysis and experimental exposures to environmentally relevant concentrations of cadmium, zinc and silver showed that G. fossarum mt1 induction is stronger after Cd exposure compared to the other tested metals. G. fossarum mt1 was more significantly induced in the caeca than in the gills of exposed organisms for any metal exposure, while G. fossarum mt2 was, at least at the individual level, more inducible in the gills than in the caeca of G. fossarum exposed to Cd and Zn. Our results provide new genetic resources that will help to improve the understanding of metal homeostasis in this sentinel species.

The CDPK Gene Family in Mustard (Brassica juncea L.): Genome-Wide Identification and Expression Analysis under Cold Stress

The CDPK family genes play crucial roles in signal transduction pathways during plant development and stress response. In this study, we comprehensively analyzed the CDPK family genes in mustard (Brassica juncea L.), resulting in the identification of 101 genes (BjuCDPK1–101) located on chromosomes AA_Chr01 to BB_Chr08. RNA-seq analysis showed that most BjuCDPK genes were predominantly expressed in the root and flower bud, suggesting their organ-specific expression patterns. Furthermore, the expression levels of most BjuCDPK genes were significantly altered by cold stress. The promoter regions of BjuCDPK genes had abundant cis-acting elements related to hormones and stress. The interaction network analysis predicted that BjuCDPK proteins respond to abiotic stress probably via interaction with ABF1, ABF4, and/or DI19. Our findings provide valuable information for further the investigation of the cold stress adaption of mustard via the CDPK signaling pathway.

Cuproptosis-related lncRNAs ovarian cancer: Multi-omics analysis of molecular mechanisms and potential therapeutic targets.

Ovarian cancer (OV) is an aggressive malignancy that poses a significant threat to the health and lives of women. Cuproptosis is a newly discovered form of programmed cell death that offers a promising therapeutic target, although its significance in cancer progression remains uncertain. In this study, we established a prognostic model of OV with six cuproptosis-related long non-coding RNAs (lncRNAs), including CTC.246B18.8, LINC00337, RP11.568N6.1, RP11.158I9.8, RP11.678G14.3 and CYP4F26P, based on the data of The Cancer Genome Atlas (TCGA). Lower risk scores were associated with favorable prognosis. In addition, a negative outcome was associated with high expression of CTC.246B18.8. According to the ESTIMATE algorithm, CTC.246B18.8 was negatively correlated with the ImmuneScore, and positively with immune checkpoints, immune cell infiltration, and tumor mutation burden (TMB). Moreover, gene set enrichment analysis (GSEA) revealed that pathways related to immunosuppression are likely activated in response to CTC-246B18.8 overexpression. Furthermore, CTC-246B18.8 expression was also associated with the sensitivity to various chemotherapy drugs. The expression patterns of the above lncRNAs were verified in ovarian tumor cell lines (SK-OV-3, COC1, and A2780) and normal ovarian epithelial cells (IOSE - 80). Six cuproptosis-related genes (CRGs), including ATP7B, MTF1, SLC31A1, DLD, ATP7A and DLAT, were differentially expressed between CTC-246B18.8high and CTC-246B18.8low patient groups, and exhibited organ-specific expression patterns pan-cancer. Small molecule drugs that target these CRGs were predicted, and potential candidates included DIAMIDE, bathocuproine disulfonate, D-penicillamine, etc. To summarize, our findings provide molecular insights into the role of cuproptosis in OV, and the signature lncRNAs and CRGs should be investigated further as immunotherapy biomarkers of OV.

Genome-wide analysis of blueberry B-box family genes and identification of members activated by abiotic stress

BackgroundB-box (BBX) proteins play important roles in regulating plant growth, development, and abiotic stress responses. BBX family genes have been identified and functionally characterized in many plant species, but little is known about the BBX family in blueberry (Vaccinium corymbosum).ResultIn this study, we identified 23 VcBBX genes from the Genome Database for Vaccinium (GDV). These VcBBXs can be divided into five clades based on gene structures and conserved domains in their encoded proteins. The prediction of cis-acting elements in the upstream sequences of VcBBX genes and protein–protein interactions indicated that VcBBX proteins are likely involved in phytohormone signaling pathways and abiotic stress responses. Analysis of transcriptome deep sequencing (RNA-seq) data showed that VcBBX genes exhibited organ-specific expression pattern and 11 VcBBX genes respond to ultraviolet B (UV-B) radiation. The co-expression analysis revealed that the encoded 11 VcBBX proteins act as bridges integrating UV-B and phytohormone signaling pathways in blueberry under UV-B radiation. Reverse-transcription quantitative PCR (RT-qPCR) analysis showed that most VcBBX genes respond to drought, salt, and cold stress. Among VcBBX proteins, VcBBX24 is highly expressed in all the organs, not only responds to abiotic stress, but it also interacts with proteins in UV-B and phytohormone signaling pathways, as revealed by computational analysis and co-expression analysis, and might be an important regulator integrating abiotic stress and phytohormone signaling networks.ConclusionsTwenty-three VcBBX genes were identified in blueberry, in which, 11 VcBBX genes respond to UV-B radiation, and act as bridges integrating UV-B and phytohormone signaling pathways according to RNA-seq data. The expression patterns under abiotic stress suggested that the functional roles of most VcBBX genes respose to drought, salt, and cold stress. Our study provides a useful reference for functional analysis of VcBBX genes and for improving abiotic stress tolerance in blueberry.

Shared and organ-specific gene-expression programs during the development of the cochlea and the superior olivary complex

ABSTRACT The peripheral and central auditory subsystems together form a complex sensory network that allows an organism to hear. The genetic programs of the two subsystems must therefore be tightly coordinated during development. Yet, their interactions and common expression pathways have never been systematically explored. MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression and are essential for normal development of the auditory system. We performed mRNA and small-RNA sequencing of organs from both auditory subsystems at three critical developmental timepoints (E16, P0, P16) to obtain a comprehensive and unbiased insight of their expression profiles. Our analysis reveals common and organ-specific expression patterns for differentially regulated mRNAs and miRNAs, which could be clustered with a particular selection of functions such as inner ear development, Wnt signalling, K+ transport, and axon guidance, based on gene ontology. Bioinformatics detected enrichment of predicted targets of specific miRNAs in the clusters and predicted regulatory interactions by monitoring opposite trends of expression of miRNAs and their targets. This approach identified six miRNAs as strong regulatory candidates for both subsystems. Among them was miR-96, an established critical factor for proper development in both subsystems, demonstrating the strength of our approach. We suggest that other miRNAs identified by this analysis are also common effectors of proper hearing acquirement. This first combined comprehensive analysis of the developmental program of the peripheral and central auditory systems provides important data and bioinformatics insights into the shared genetic program of the two sensory subsystems and their regulation by miRNAs.

Locational memory of macrovessel vascular cells is transcriptionally imprinted

Vascular pathologies show locational predisposition throughout the body; further insights into the transcriptomics basis of this vascular heterogeneity are needed. We analyzed transcriptomes from cultured endothelial cells and vascular smooth muscle cells from nine adult canine macrovessels: the aorta, coronary artery, vena cava, portal vein, femoral artery, femoral vein, saphenous vein, pulmonary vein, and pulmonary artery. We observed that organ-specific expression patterns persist in vitro, indicating that these genes are not regulated by blood flow or surrounding cell types but are likely fixed in the epigenetic memory. We further demonstrated the preserved location-specific expression of GATA4 protein in cultured cells and in the primary adult vessel. On a functional level, arterial and venous endothelial cells differed in vascular network morphology as the arterial networks maintained a higher complexity. Our findings prompt the rethinking of the extrapolation of results from single-origin endothelial cell systems.

Of tenants and nomads: The faces of memory T cells

Memory Tcells comprise circulating and tissue-resident subsets. In this issue of Immunity, Evrard etal. generate an imputed high-dimensional, single-cell protein expression atlas of memory CD8+ Tcells, providing insights into stable differentiation markers and organ-specific expression patterns.

Identification and functional characterization of three cytochrome P450 genes for the abietane diterpenoid biosynthesis in Isodon lophanthoides

We identify two ferruginol synthases and a 11-hydroxyferruginol synthase from a traditional Chinese medicinal herb Isodon lophanthoides and propose their involvement in two independent abietane diterpenoids biosynthetic pathways. Isodon lophanthoides is a traditional Chinese medicinal herb rich in highly oxidized abietane-type diterpenoids. These compounds exhibit a wide range of pharmaceutical activities, yet the biosynthesis is barely known. Here, we describe the screening and functional characterization of P450s that oxidize the abietane skeleton abietatriene. We mainly focused on CYP76 family and identified 12 CYP76AHs by mining the RNA-seq data of I. lophanthoides. Among the 12 CYP76AHs, 6 exhibited similar transcriptional expression features as upstream diterpene synthases, including root or leaf-preferential expression pattern and highly MeJA inducibility. These six P450s were considered as first-tier candidates and functionally characterized in yeast and plant cells. In yeast assays showed that both CYP76AH42 and CYP76AH43 were ferruginol synthases hydroxylating the C12 position of abietatriene, whereas CYP76AH46 was characterized as a 11-hydroxyferruginol synthase whichcatalyzes two successive oxidations at C12 and C11 of abietatriene. Heterologous expression of three CYP76AHs in Nicotiana benthamiana resulted in the formation of ferruginol. qPCR analysis showed CYP76AH42 and CYP76AH43 were mainly expressed in the root, which was consistent with the distribution of ferruginol in the root periderms. CYP76AH46 was primarily expressed in the leaves where barely ferruginol or 11-hydroxyferruginol was detected. In addition to distinct organ-specific expression pattern, three CYP76AHs exhibited different genomic structures (w or w/o introns), low protein sequence identities (51-63%) and were placed in separate subclades in the phylogenetic tree. These results suggest that the identified CYP76AHs may be involved in at least two independent abietane biosynthetic pathways in the aerial and underground parts of I. lophanthoides.

The combination of SMRT sequencing and Illumina sequencing highlights organ-specific and age-specific expression patterns of miRNAs in Sika Deer.

Due to the lack of high-quality Sika Deer (Cervus nippon) transcriptome and sRNAome across multiple organs or development stages, it is impossible to comprehensively analyze the mRNA and miRNA regulatory networks related to growth, development and immunity response. In this study, we used single molecule-real time sequencing (SMRT-seq) and Illumina sequencing methods to generate transcriptome and sRNAome from ten tissues and four age groups of Sika Deer to help us understand molecular characteristics and global miRNA expression profiles. The results showed that a total of 240,846 consensus transcripts were generated with an average length of 2,784 bp. 4,329 Transcription factors (TFs), 109,000 Simple Sequence Repeats (SSRs) and 18,987 Long non-coding RNAs (LncRNAs) were identified. Meanwhile, 306 known miRNAs and 143 novel miRNAs were obtained. A large number of miRNAs showed organ-specific and age-specific differential expression patterns. In particular, we found that the organ-specific miRNAs were enriched in the brain, some of which shared only between the brain and adrenal. These miRNAs were involved in maintaining specific functions within the brain and adrenal. By constructing miRNA96mRNA interaction networks associated with Sika Deer immunity, we found that miRNAs (miR-148a, miR-26a, miR-214, let-7b, etc.) and mRNAs (CD6, TRIM38, C3, CD163, etc.) might play an important role in the immune response of Sika Deer spleen. Together, our study generated an improved transcript annotation for Sika Deer by SMRT-seq and revealed the role of miRNA in regulating the growth, development and immunity response of Sika Deer.

Genome-Wide Identification of the Highly Conserved INDETERMINATE DOMAIN (IDD) Zinc Finger Gene Family in Moso Bamboo (Phyllostachys edulis).

INDETERMINATE DOMAIN (IDD) proteins, a family of transcription factors unique to plants, function in multiple developmental processes. Although the IDD gene family has been identified in many plants, little is known about it in moso bamboo. In this present study, we identified 32 PheIDD family genes in moso bamboo and randomly sequenced the full-length open reading frames (ORFs) of ten PheIDDs. All PheIDDs shared a highly conserved IDD domain that contained two canonical C2H2-ZFs, two C2HC-ZFs, and a nuclear localization signal. Collinearity analysis showed that segmental duplication events played an important role in expansion of the PheIDD gene family. Synteny analysis indicated that 30 PheIDD genes were orthologous to those of rice (Oryza sativa). Thirty PheIDDs were expressed at low levels, and most PheIDDs exhibited characteristic organ-specific expression patterns. Despite their diverse expression patterns in response to exogenous plant hormones, 8 and 22 PheIDDs responded rapidly to IAA and 6-BA treatments, respectively. The expression levels of 23 PheIDDs were closely related to the outgrowth of aboveground branches and 20 PheIDDs were closely related to the awakening of underground dormant buds. In addition, we found that the PheIDD21 gene generated two products by alternative splicing. Both isoforms interacted with PheDELLA and PheSCL3. Furthermore, both isoforms could bind to the cis-elements of three genes (PH02Gene17121, PH02Gene35441, PH02Gene11386). Taken together, our work provides valuable information for studying the molecular breeding mechanism of lateral organ development in moso bamboo.

Genome-Wide Identification and Expression Analysis of Eggplant DIR Gene Family in Response to Biotic and Abiotic Stresses

Dirigent proteins (DIR) play important roles in the biosynthesis of lignins and lignans, defensive responses, secondary metabolism, and disease resistance in plants. The DIR gene family has been identified and studied in many plants. However, the identification of DIR gene family in eggplant has not been conducted yet. Therefore, in this study, based on the available genome information of eggplant, the DIR family genes in eggplant were identified with bioinformatics methods. The expression pattern analyses of eggplant DIR family genes in different organs and stresses were also conducted to understand their biological functions. The results showed that a total of 24 DIR genes were identified in the eggplant, which were divided into three subfamilies (DIR-a, DIR-b/d, and DIR-e). Synteny analysis of DIR genes among eggplant, Arabidopsis, and rice showed that 15 eggplant DIR genes were colinear with 18 Arabidopsis DIR genes, and 16 eggplant DIR genes were colinear with 15 rice DIR genes. Phylogenetic tree analysis showed that 19 pairs of orthologous genes were identified between eggplant and pepper. The cis-acting elements analysis implied that the eggplant DIR genes contained a lot of cis-elements associated with stress and hormone response. The organ-specific expression analysis of eggplant DIR family genes revealed that only the SmDIR3 gene was highly expressed in all the 19 organs of eggplant. Some SmDIR genes, including SmDIR7, SmDIR8, SmDIR11, SmDIR14, SmDIR18, SmDIR19, SmDIR20, and SmDIR23, were not or were lowly expressed in the eggplant organs, while the other eggplant DIR family genes showed an organ-specific expression pattern. Furthermore, 19 of 24 SmDIR genes were differentially expressed in response to abiotic and biotic stresses. 5 SmDIR genes, including SmDIR3, SmDIR5, SmDIR6, SmDIR12, and SmDIR22, were differentially expressed under multiple types of abiotic and biotic stresses. Especially notable, the SmDIR22 gene was differentially expressed under three types of abiotic stresses and two types of biotic stresses, which indicated that the SmDIR22 gene plays an important role in the response to abiotic and biotic stresses. These results provide valuable evidence for a better understanding of the biological role of DIR genes in eggplant.

Evolution and functional diversification of catalase genes in the green lineage

BackgroundCatalases (CATs) break down hydrogen peroxide into water and oxygen to prevent cellular oxidative damage, and play key roles in the development, biotic and abiotic stresses of plants. However, the evolutionary relationships of the plant CAT gene family have not been systematically reported.ResultsHere, we conducted genome-wide comparative, phylogenetic, and structural analyses of CAT orthologs from 29 out of 31 representative green lineage species to characterize the evolution and functional diversity of CATs. We found that CAT genes in land plants were derived from core chlorophytes and detected a lineage-specific loss of CAT genes in Fabaceae, suggesting that the CAT genes in this group possess divergent functions. All CAT genes were split into three major groups (group α, β1, and β2) based on the phylogeny. CAT genes were transferred from bacteria to core chlorophytes and charophytes by lateral gene transfer, and this led to the independent evolution of two types of CAT genes: α and β types. Ten common motifs were detected in both α and β groups, and β CAT genes had five unique motifs, respectively. The findings of our study are inconsistent with two previous hypotheses proposing that (i) new CAT genes are acquired through intron loss and that (ii) the Cys-343 residue is highly conserved in plants. We found that new CAT genes in most higher plants were produced through intron acquisition and that the Cys-343 residue was only present in monocots, Brassicaceae and Pp_CatX7 in P. patens, which indicates the functional specificity of the CATs in these three lineages. Finally, our finding that CAT genes show high overall sequence identity but that individual CAT genes showed developmental stage and organ-specific expression patterns suggests that CAT genes have functionally diverged independently.ConclusionsOverall, our analyses of the CAT gene family provide new insights into their evolution and functional diversification in green lineage species.

An organ-specific transcriptomic atlas of the medicinal plant Bletilla striata: Protein-coding genes, microRNAs, and regulatory networks.

As one of the important species belonging to the Bletilla genus of Orchidaceae, Bletilla striata (Thunb.) Rchb. f., possesses both ornamental and medicinal values. Its dried tubers are used as a traditional Chinese medicine, and several secondary metabolites have been indicated to be the active ingredients. However, the molecular mechanisms related to the regulation of secondary metabolism have not been characterized in B. striata. In this study, integrated analysis of RNA sequencing (RNA-seq), small RNA sequencing (sRNA-seq), and degradome sequencing (degradome-seq) data from three organs (leaf, root, and tuber) of B. striata provided us with a comprehensive view of the microRNA (miRNA)-mediated regulatory network. Firstly, based on the RNA-seq data, the organ-specific expression patterns of the protein-coding genes, especially for those related to secondary metabolism, were investigated. Secondly, 342 conserved miRNA candidates were identified from B. striata. These miRNAs were assigned to 88 families, some of which were selected for expression pattern analysis. Additionally, 31 hairpin-structured precursors encoding 23 novel miRNAs were uncovered from the transcriptome assembly. Thirdly, based on the degradome signatures, 1,142 validated miRNA-target pairs (involving 167 conserved miRNAs and six novel miRNAs and 51 target genes) were included in the regulatory network. Organ-specific expression level comparison between the miRNAs and their targets revealed some interesting miRNA-target pairs. Fourthly, some valuable subnetworks were extracted for further functional studies. Additionally, some regulatory pathways were indicated to be monocot specific. Summarily, our results lay a solid basis for in-depth studies on the regulatory mechanisms underlying the production of the medicinal ingredients in B. striata.

Profiles of Cytokinins Metabolic Genes and Endogenous Cytokinins Dynamics during Shoot Multiplication In Vitro of Phalaenopsis.

Shoot multiplication induced by exogenous cytokinins (CKs) has been commonly used in Phalaenopsis micropropagation for commercial production. Despite this, mechanisms of CKs action on shoot multiplication remain unclear in Phalaenopsis. In this study, we first identified key CKs metabolic genes, including six isopentenyltransferase (PaIPTs), six cytokinin riboside 5′ monophosphate phosphoribohydrolase (PaLOGs), and six cytokinin dehydrogenase (PaCKXs), from the Phalaenopsis genome. Then, we investigated expression profiles of these CKs metabolic genes and endogenous CKs dynamics in shoot proliferation by thidiazuron (TDZ) treatments (an artificial plant growth regulator with strong cytokinin-like activity). Our data showed that these CKs metabolic genes have organ-specific expression patterns. The shoot proliferation in vitro was effectively promoted with increased TDZ concentrations. Following TDZ treatments, the highly expressed CKs metabolic genes in micropropagated shoots were PaIPT1, PaLOG2, and PaCKX4. By 30 days of culture, TDZ treatments significantly induced CK-ribosides levels in micropropagated shoots, such as tZR and iPR (2000-fold and 200-fold, respectively) as compared to the controls, whereas cZR showed only a 10-fold increase. Overexpression of PaIPT1 and PaLOG2 by agroinfiltration assays resulted in increased CK-ribosides levels in tobacco leaves, while overexpression of PaCKX4 resulted in decreased CK-ribosides levels. These findings suggest de novo biosynthesis of CKs induced by TDZ, primarily in elevation of tZR and iPR levels. Our results provide a better understanding of CKs metabolism in Phalaenopsis micropropagation.

Identification of macrophages in normal and injured mouse tissues using reporter lines and antibodies

Reliable tools for macrophage identification in mouse tissues are critical for studies investigating inflammatory and reparative responses. Transgenic reporter mice and anti-macrophage antibodies have been used as “specific pan-macrophage” markers in many studies; however, organ-specific patterns of expression and non-specific labeling of other cell types, such as fibroblasts, may limit their usefulness. Our study provides a systematic comparison of macrophage labeling patterns in normal and injured mouse tissues, using the CX3CR1 and CSF1R macrophage reporter lines and anti-macrophage antibodies. Moreover, we tested the specificity of macrophage antibodies using the fibroblast-specific PDGFRmathrm{alpha } reporter line. Mouse macrophages exhibit organ-specific differences in expression of macrophage markers. Hepatic macrophages are labeled for CSF1R, Mac2 and F4/80, but lack CX3CR1 expression, whereas in the lung, the CSF1R+/Mac2+/Mac3+ macrophage population is not labeled with F4/80. In the splenic red pulp, subpopulations of CSF1R+/F4/80+/Mac3+cells were labeled with Mac2, CX3CR1 and lysozyme M. In the kidney, Mac2, Mac3 and lysozyme M labeled a fraction of the CSF1R+ and CX3CR1+ macrophages, but also stained tubular epithelial cells. In normal hearts, the majority of CSF1R+ and CX3CR1+ cells were not detected with anti-macrophage antibodies. Myocardial infarction was associated with marked expansion of the CSF1R+ and CX3CR1+ populations that peaked during the proliferative phase of cardiac repair, and also expressed Mac2, Mac3 and lysozyme M. In normal mouse tissues, a small fraction of cells labeled with anti-macrophage antibodies were identified as PDGFRmathrm{alpha }+ fibroblasts, using a reporter system. The population of PDGFRmathrm{alpha }+ cells expressing macrophage markers expanded following injury, likely reflecting emergence of cellular phenotypes with both fibroblast and macrophage characteristics. In conclusion, mouse macrophages exhibit remarkable heterogeneity. Selection of the most appropriate markers for identification of macrophages in mouse tissues is dependent on the organ and the pathologic condition studied.

Tomato SlCER1-1 catalyzes the synthesis of wax alkanes which increases the drought tolerance and fruit storability.

Very-long-chain (VLC) alkanes are the main wax compounds of tomato fruits and leaves. ECERIFERUM1 (CER1) and ECERIFERUM3 (CER3) are the two key genes involved in VLC alkane biosynthesis in Arabidopsis thaliana. However, CER1 and CER3 homologs have not been investigated in tomato, and their exact biological functions remain unknown. We analyzed the wax profiles of tomato leaves and fruits at different growth stages and characterized the tomato CER1 and CER3 homologs. VLC alkanes were the predominant wax compounds in both leaves and fruits at all developmental stages. We identified five CER1 homologs and two CER3 homologs in tomato, which were designated SlCER1–1 to SlCER1–5 and SlCER3–1 and SlCER3–2, respectively. The genes exhibited tissue- and organ-specific expression patterns and were induced by abiotic stresses. SlCER1–1 was localized to the endoplasmic reticulum (ER), which is also the main site of wax biosynthesis. Silencing SlCER1–1 in tomato significantly reduced the amounts of n-alkanes and branched alkanes, whereas its overexpression in Arabidopsis had the opposite effect. Under drought stress, both n-alkanes and branched alkanes increased significantly in wild-type but not SlCER1–1 RNAi tomato plants. Furthermore, SlCER1–1 silencing also increased the cuticular permeability of leaves and fruits. In conclusion, SlCER1–1 is involved in wax alkane biosynthesis in tomato and plays an important role in drought tolerance and fruit storability.