Common Cis-regulatory Elements Research Articles

Epigenetic modifications and miRNAs determine the transition of somatic cells into somatic embryos.

This review discusses the epigenetic changes during somatic embryo (SE) development, highlights the genes and miRNAs involved in the transition of somatic cells into SEs as a result of epigenetic changes, and draws insights on biotechnological opportunities to study SE development. Somatic embryogenesis from somatic cells occurs in a series of steps. The transition of somatic cells into somatic embryos (SEs) is the most critical step under genetic and epigenetic regulations. Major regulatory genes such as SERK, WUS, BBM, FUS3/FUSA3, AGL15, and PKL, control SE steps and development by turning on and off other regulatory genes. Gene transcription profiles of somatic cells during SE development is the result of epigenetic changes, such as DNA and histone protein modifications, that control and decide the fate of SE formation. Depending on the type of somatic cells and the treatment with plant growth regulators, epigenetic changes take place dynamically. Either hypermethylation or hypomethylation of SE-related genes promotes the transition of somatic cells. For example, the reduced levels of DNA methylation of SERK and WUS promotes SE initiation. Histone modifications also promote SE induction by regulating SE-related genes in somatic cells. In addition, miRNAs contribute to the various stages of SE by regulating the expression of auxin signaling pathway genes (TIR1, AFB2, ARF6, and ARF8), transcription factors (CUC1 and CUC2), and growth-regulating factors (GRFs) involved in SE formation. These epigenetic and miRNA functions are unique and have the potential to regenerate bipolar structures from somatic cells when a pluripotent state is induced. However, an integrated overview of the key regulators involved in SE development and downstream processes is lacking. Therefore, this review discusses epigenetic modifications involved in SE development, SE-related genes and miRNAs associated with epigenetics, and common cis-regulatory elements in the promoters of SE-related genes. Finally, we highlight future biotechnological opportunities to alter epigenetic pathways using the genome editing tool and to study the transition mechanism of somatic cells.

Targeting cis-regulatory elements of FOXO family is a novel therapeutic strategy for induction of leukemia cell differentiation

Differentiation therapy has been proposed as a promising therapeutic strategy for acute myeloid leukemia (AML); thus, the development of more versatile methodologies that are applicable to a wide range of AML subtypes is desired. Although the FOXOs transcription factor represents a promising drug target for differentiation therapy, the efficacy of FOXO inhibitors is limited in vivo. Here, we show that pharmacological inhibition of a common cis-regulatory element of forkhead box O (FOXO) family members successfully induced cell differentiation in various AML cell lines. Through gene expression profiling and differentiation marker-based CRISPR/Cas9 screening, we identified TRIB1, a complement of the COP1 ubiquitin ligase complex, as a functional FOXO downstream gene maintaining an undifferentiated status. TRIB1 is direct target of FOXO3 and the FOXO-binding cis-regulatory element in the TRIB1 promoter, referred to as the FOXO-responsive element in the TRIB1 promoter (FRE-T), played a critical role in differentiation blockade. Thus, we designed a DNA-binding pharmacological inhibitor of the FOXO-FRE-T interface using pyrrole-imidazole polyamides (PIPs) that specifically bind to FRE-T (FRE-PIPs). The FRE-PIPs conjugated to chlorambucil (FRE-chb) inhibited transcription of TRIB1, causing differentiation in various AML cell lines. FRE-chb suppressed the formation of colonies derived from AML cell lines but not from normal counterparts. Administration of FRE-chb inhibited tumor progression in vivo without remarkable adverse effects. In conclusion, targeting cis-regulatory elements of the FOXO family is a promising therapeutic strategy that induces AML cell differentiation.

Trans-species microRNA loci in the parasitic plant Cuscuta campestris have a U6-like snRNA promoter.

Small regulatory RNAs can move between organisms and regulate gene expression in the recipient. Whether the trans-species small RNAs being exported are distinguished from the normal endogenous small RNAs of the source organism is not known. The parasitic plant Cuscuta campestris (dodder) produces many microRNAs that specifically accumulate at the host-parasite interface, several of which have trans-species activity. We found that induction of C. campestris interface-induced microRNAs is similar regardless of host species and occurs in C. campestris haustoria produced in the absence of any host. The loci-encoding C. campestris interface-induced microRNAs are distinguished by a common cis-regulatory element. This element is identical to a conserved upstream sequence element (USE) used by plant small nuclear RNA loci. The properties of the interface-induced microRNA primary transcripts strongly suggest that they are produced via U6-like transcription by RNA polymerase III. The USE promotes accumulation of interface-induced miRNAs (IIMs) in a heterologous system. This promoter element distinguishes C. campestris IIM loci from other plant small RNAs. Our data suggest that C. campestris IIMs are produced in a manner distinct from canonical miRNAs. All confirmed C. campestris microRNAs with documented trans-species activity are interface-induced and possess these features. We speculate that RNA polymerase III transcription of IIMs may allow these miRNAs to be exported to hosts.

CRISPR/Cas9 screen uncovers functional translation of cryptic lncRNA-encoded open reading frames in human cancer.

Emerging evidence suggests that cryptic translation within long noncoding RNAs (lncRNAs) may produce novel proteins with important developmental/physiological functions. However, the role of this cryptic translation in complex diseases (e.g., cancer) remains elusive. Here, we applied an integrative strategy combining ribosome profiling and CRISPR/Cas9 screening with large-scale analysis of molecular/clinical data for breast cancer (BC) and identified estrogen receptor α-positive (ER+) BC dependency on the cryptic ORFs encoded by lncRNA genes that were upregulated in luminal tumors. We confirmed the in vivo tumor-promoting function of an unannotated protein, GATA3-interacting cryptic protein (GT3-INCP) encoded by LINC00992, the expression of which was associated with poor prognosis in luminal tumors. GTE-INCP was upregulated by estrogen/ER and regulated estrogen-dependent cell growth. Mechanistically, GT3-INCP interacted with GATA3, a master transcription factor key to mammary gland development/BC cell proliferation, and coregulated a gene expression program that involved many BC susceptibility/risk genes and impacted estrogen response/cell proliferation. GT3-INCP/GATA3 bound to common cis regulatory elements and upregulated the expression of the tumor-promoting and estrogen-regulated BC susceptibility/risk genes MYB and PDZK1. Our study indicates that cryptic lncRNA-encoded proteins can be an important integrated component of the master transcriptional regulatory network driving aberrant transcription in cancer, and suggests that the "hidden" lncRNA-encoded proteome might be a new space for therapeutic target discovery.

Characterization of promoter elements of isoprene-responsive genes and the ability of isoprene to bind START domain transcription factors.

Isoprene has recently been proposed to be a signaling molecule that can enhance tolerance of both biotic and abiotic stress. Not all plants make isoprene, but all plants tested to date respond to isoprene. We hypothesized that isoprene interacts with existing signaling pathways rather than requiring novel mechanisms for its effect on plants. We analyzed the cis-regulatory elements (CREs) in promoters of isoprene-responsive genes and the corresponding transcription factors binding these promoter elements to obtain clues about the transcription factors and other proteins involved in isoprene signaling. Promoter regions of isoprene-responsive genes were characterized using the Arabidopsis cis-regulatory element database. CREs bind ARR1, Dof, DPBF, bHLH112, GATA factors, GT-1, MYB, and WRKY transcription factors, and light-responsive elements were overrepresented in promoters of isoprene-responsive genes; CBF-, HSF-, WUS-binding motifs were underrepresented. Transcription factors corresponding to CREs overrepresented in promoters of isoprene-responsive genes were mainly those important for stress responses: drought-, salt/osmotic-, oxidative-, herbivory/wounding and pathogen-stress. More than half of the isoprene-responsive genes contained at least one binding site for TFs of the class IV (homeodomain leucine zipper) HD-ZIP family, such as GL2, ATML1, PDF2, HDG11, ATHB17. While the HD-zipper-loop-zipper (ZLZ) domain binds to the L1 box of the promoter region, a special domain called the steroidogenic acute regulatory protein-related lipid transfer, or START domain, can bind ligands such as fatty acids (e.g., linolenic and linoleic acid). We tested whether isoprene might bind in such a START domain. Molecular simulations and modeling to test interactions between isoprene and a class IV HD-ZIP family START-domain-containing protein were carried out. Without membrane penetration by the HDG11 START domain, isoprene within the lipid bilayer was inaccessible to this domain, preventing protein interactions with membrane bound isoprene. The cross-talk between isoprene-mediated signaling and other growth regulator and stress signaling pathways, in terms of common CREs and transcription factors could enhance the stability of the isoprene emission trait when it evolves in a plant but so far it has not been possible to say what how isoprene is sensed to initiate signaling responses.

A genome-wide association study to identify candidate genes for erectile dysfunction.

Erectile dysfunction (ED) can be caused by different diseases and controlled by several genetic networks. In this study, to identify the genes related to ED, the expression profiles of normal and ED samples were investigated by the Gene Expression Omnibus (GEO) database. Seventeen genes were identified as associated genes with ED. The protein and nucleic acid sequences of selected genes were retrieved from the UCSC database. Selected genes were diverse according to their physicochemical properties and functions. Category function revealed that selected genes are involved in pathways related to humans some diseases. Furthermore, based on protein interactions, genes associated with the insulin pathway had the greatest interaction with the studied genes. To identify the common cis-regulatory elements, the promoter site of the selected genes was retrieved from the UCSC database. The Gapped Local Alignment of Motifs tool was used for finding common conserved motifs into the promoter site of selected genes. Besides, INSR protein as an insulin receptor precursor showed a high potential site for posttranslation modifications, including phosphorylation and N-glycosylation. Also, in this study, two Guanine-Cytosine (GC)-rich regions were identified as conserved motifs in the upstream of studied genes which can be involved in regulating the expression of genes associated with ED. Also, the conserved binding site of miR-29-3p that is involved in various cancers was observed in the 3' untranslated region of genes associated with ED. Our study introduced new genes associated with ED, which can be good candidates for further analyzing related to human ED.

Structural basis for the recognition of transiently structured AU-rich elements by Roquin.

Adenylate/uridylate-rich elements (AREs) are the most common cis-regulatory elements in the 3′-untranslated region (UTR) of mRNAs, where they fine-tune turnover by mediating mRNA decay. They increase plasticity and efficacy of mRNA regulation and are recognized by several ARE-specific RNA-binding proteins (RBPs). Typically, AREs are short linear motifs with a high content of complementary A and U nucleotides and often occur in multiple copies. Although thermodynamically rather unstable, the high AU-content might enable transient secondary structure formation and modify mRNA regulation by RBPs. We have recently suggested that the immunoregulatory RBP Roquin recognizes folded AREs as constitutive decay elements (CDEs), resulting in shape-specific ARE-mediated mRNA degradation. However, the structural evidence for a CDE-like recognition of AREs by Roquin is still lacking. We here present structures of CDE-like folded AREs, both in their free and protein-bound form. Moreover, the AREs in the UCP3 3′-UTR are additionally bound by the canonical ARE-binding protein AUF1 in their linear form, adopting an alternative binding-interface compared to the recognition of their CDE structure by Roquin. Strikingly, our findings thus suggest that AREs can be recognized in multiple ways, allowing control over mRNA regulation by adapting distinct conformational states, thus providing differential accessibility to regulatory RBPs.

Is there an aesthetics in golden ratio as regards to the common cis-regulatory elements versus to atomic numbers of elements with respect to Quantum perspective model?

Is there an aesthetics in golden ratio as regards to the common cis-regulatory elements versus to atomic numbers of elements with respect to Quantum perspective model?

Conserved Gene Microsynteny Unveils Functional Interaction Between Protein Disulfide Isomerase and Rho Guanine-Dissociation Inhibitor Families

Protein disulfide isomerases (PDIs) support endoplasmic reticulum redox protein folding and cell-surface thiol-redox control of thrombosis and vascular remodeling. The family prototype PDIA1 regulates NADPH oxidase signaling and cytoskeleton organization, however the related underlying mechanisms are unclear. Here we show that genes encoding human PDIA1 and its two paralogs PDIA8 and PDIA2 are each flanked by genes encoding Rho guanine-dissociation inhibitors (GDI), known regulators of RhoGTPases/cytoskeleton. Evolutionary histories of these three microsyntenic regions reveal their emergence by two successive duplication events of a primordial gene pair in the last common vertebrate ancestor. The arrangement, however, is substantially older, detectable in echinoderms, nematodes, and cnidarians. Thus, PDI/RhoGDI pairing in the same transcription orientation emerged early in animal evolution and has been largely maintained. PDI/RhoGDI pairs are embedded into conserved genomic regions displaying common cis-regulatory elements. Analysis of gene expression datasets supports evidence for PDI/RhoGDI coexpression in developmental/inflammatory contexts. PDIA1/RhoGDIα were co-induced in endothelial cells upon CRISP-R-promoted transcription activation of each pair component, and also in mouse arterial intima during flow-induced remodeling. We provide evidence for physical interaction between both proteins. These data support strong functional links between PDI and RhoGDI families, which likely maintained PDI/RhoGDI microsynteny along > 800-million years of evolution.

The Eucalyptus Tonoplast Intrinsic Protein (TIP) Gene Subfamily: Genomic Organization, Structural Features, and Expression Profiles.

Plant aquaporins are water channels implicated in various physiological processes, including growth, development and adaptation to stress. In this study, the Tonoplast Intrinsic Protein (TIP) gene subfamily of Eucalyptus, an economically important woody species, was investigated and characterized. A genome-wide survey of the Eucalyptus grandis genome revealed the presence of eleven putative TIP genes (referred as EgTIP), which were individually assigned by phylogeny to each of the classical TIP1–5 groups. Homology modeling confirmed the presence of the two highly conserved NPA (Asn-Pro-Ala) motifs in the identified EgTIPs. Residue variations in the corresponding selectivity filters, that might reflect differences in EgTIP substrate specificity, were observed. All EgTIP genes, except EgTIP5.1, were transcribed and the majority of them showed organ/tissue-enriched expression. Inspection of the EgTIP promoters revealed the presence of common cis-regulatory elements implicated in abiotic stress and hormone responses pointing to an involvement of the identified genes in abiotic stress responses. In line with these observations, additional gene expression profiling demonstrated increased expression under polyethylene glycol-imposed osmotic stress. Overall, the results obtained suggest that these novel EgTIPs might be functionally implicated in eucalyptus adaptation to stress.

Discriminative gene co-expression network analysis uncovers novel modules involved in the formation of phosphate deficiency-induced root hairs in Arabidopsis.

Cell fate and differentiation in the Arabidopsis root epidermis are genetically defined but remain plastic to environmental signals such as limited availability of inorganic phosphate (Pi). Root hairs of Pi-deficient plants are more frequent and longer than those of plants grown under Pi-replete conditions. To dissect genes involved in Pi deficiency-induced root hair morphogenesis, we constructed a co-expression network of Pi-responsive genes against a customized database that was assembled from experiments in which differentially expressed genes that encode proteins with validated functions in root hair development were over-represented. To further filter out less relevant genes, we combined this procedure with a search for common cis-regulatory elements in the promoters of the selected genes. In addition to well-described players and processes such as auxin signalling and modifications of primary cell walls, we discovered several novel aspects in the biology of root hairs induced by Pi deficiency, including cell cycle control, putative plastid-to-nucleus signalling, pathogen defence, reprogramming of cell wall-related carbohydrate metabolism, and chromatin remodelling. This approach allows the discovery of novel of aspects of a biological process from transcriptional profiles with high sensitivity and accuracy.

Analysis of cis-regulatory element distribution in gene promoters of Gossypium raimondii and Arabidopsis thaliana

Cotton genomic studies have boomed since the release of Gossypium raimondii draft genome. In this study, cis-regulatory element (CRE) in 1 kb length sequence upstream 5' UTR of annotated genes were selected and scanned in the Arabidopsis thaliana (At) and Gossypium raimondii (Gr) genomes, based on the database of PLACE (Plant cis-acting Regulatory DNA Elements). According to the definition of this study, 44 (12.3%) and 57 (15.5%) CREs presented "peak-like" distribution in the 1 kb selected sequences of both genomes, respectively. Thirty-four of them were peak-like distributed in both genomes, which could be further categorized into 4 types based on their core sequences. The coincidence of TATABOX peak position and their actual position ((-) -30 bp) indicated that the position of a common CRE was conservative in different genes, which suggested that the peak position of these CREs was their possible actual position of transcription factors. The position of a common CRE was also different between the two genomes due to stronger length variation of 5' UTR in Gr than At. Furthermore, most of the peak-like CREs were located in the region of -110 bp-0 bp, which suggested that concentrated distribution might be conductive to the interaction of transcription factors, and then regulate the gene expression in downstream.

Genome-wide analysis of 3′-untranslated regions supports the existence of post-transcriptional regulons controlling gene expression in trypanosomes

In eukaryotic cells, a group of messenger ribonucleic acids (mRNAs) encoding functionally interrelated proteins together with the trans-acting factors that coordinately modulate their expression is termed a post-transcriptional regulon, due to their partial analogy to a prokaryotic polycistron. This mRNA clustering is organized by sequence-specific RNA-binding proteins (RBPs) that bind cis-regulatory elements in the noncoding regions of genes, and mediates the synchronized control of their fate. These recognition motifs are often characterized by conserved sequences and/or RNA structures, and it is likely that various classes of cis-elements remain undiscovered. Current evidence suggests that RNA regulons govern gene expression in trypanosomes, unicellular parasites which mainly use post-transcriptional mechanisms to control protein synthesis. In this study, we used motif discovery tools to test whether groups of functionally related trypanosomatid genes contain a common cis-regulatory element. We obtained conserved structured RNA motifs statistically enriched in the noncoding region of 38 out of 53 groups of metabolically related transcripts in comparison with a random control. These motifs have a hairpin loop structure, a preferred sense orientation and are located in close proximity to the open reading frames. We found that 15 out of these 38 groups represent unique motifs in which most 3′-UTR signature elements were group-specific. Two extensively studied Trypanosoma cruzi RBPs, TcUBP1 and TcRBP3 were found associated with a few candidate RNA regulons. Interestingly, 13 motifs showed a strong correlation with clusters of developmentally co-expressed genes and six RNA elements were enriched in gene clusters affected after hyperosmotic stress. Here we report a systematic genome-wide in silico screen to search for novel RNA-binding sites in transcripts, and describe an organized network of several coordinately regulated cohorts of mRNAs in T. cruzi. Moreover, we found that structured RNA elements are also conserved in other human pathogens. These results support a model of regulation of gene expression by multiple post-transcriptional regulons in trypanosomes.

Comparative Genomics Identifies the Mouse Bmp3 Promoter and an Upstream Evolutionary Conserved Region (ECR) in Mammals

The Bone Morphogenetic Protein (BMP) pathway is a multi-member signaling cascade whose basic components are found in all animals. One member, BMP3, which arose more recently in evolution and is found only in deuterostomes, serves a unique role as an antagonist to both the canonical BMP and Activin pathways. However, the mechanisms that control BMP3 expression, and the cis-regulatory regions mediating this regulation, remain poorly defined. With this in mind, we sought to identify the Bmp3 promoter in mouse (M. musculus) through functional and comparative genomic analyses. We found that the minimal promoter required for expression in resides within 0.8 kb upstream of Bmp3 in a region that is highly conserved with rat (R. norvegicus). We also found that an upstream region abutting the minimal promoter acts as a repressor of the minimal promoter in HEK293T cells and osteoblasts. Strikingly, a portion of this region is conserved among all available eutherian mammal genomes (47/47), but not in any non-eutherian animal (0/136). We also identified multiple conserved transcription factor binding sites in the Bmp3 upstream ECR, suggesting that this region may preserve common cis-regulatory elements that govern Bmp3 expression across eutherian mammals. Since dysregulation of BMP signaling appears to play a role in human health and disease, our findings may have application in the development of novel therapeutics aimed at modulating BMP signaling in humans.

Common SNP-Based Haplotype Analysis of the 4p16.3 Huntington Disease Gene Region

Age at the onset of motor symptoms in Huntington disease (HD) is determined largely by the length of a CAG repeat expansion in HTT but is also influenced by other genetic factors. We tested whether common genetic variation near the mutation site is associated with differences in the distribution of expanded CAG alleles or age at the onset of motor symptoms. To define disease-associated single-nucleotide polymorphisms (SNPs), we compared 4p16.3 SNPs in HD subjects with population controls in a case:control strategy, which revealed that the strongest signals occurred at a great distance from the HD mutation as a result of "synthetic association" with SNP alleles that are of low frequency in population controls. Detailed analysis delineated a prominent ancestral haplotype that accounted for ∼50% of HD chromosomes and extended to at least 938 kb on about half of these. Together, the seven most abundant haplotypes accounted for ∼83% of HD chromosomes. Neither the extended shared haplotype nor the individual local HTT haplotypes were associated with altered CAG-repeat length distribution or residual age at the onset of motor symptoms, arguing against modification of these disease features by common cis-regulatory elements. Similarly, the 11 most frequent control haplotypes showed no trans-modifier effect on age at the onset of motor symptoms. Our results argue against common local regulatory variation as a factor influencing HD pathogenesis, suggesting that genetic modifiers be sought elsewhere in the genome. They also indicate that genome-wide association analysis with a small number of cases can be effective for regional localization of genetic defects, even when a founder effect accounts for only a fraction of the disorder.

Identification of the Cluster Control Region for the Protocadherin-β Genes Located beyond the Protocadherin-γ Cluster

The clustered protocadherins (Pcdhs), Pcdh-α, -β, and -γ, are transmembrane proteins constituting a subgroup of the cadherin superfamily. Each Pcdh cluster is arranged in tandem on the same chromosome. Each of the three Pcdh clusters shows stochastic and combinatorial expression in individual neurons, thus generating a hugely diverse set of possible cell surface molecules. Therefore, the clustered Pcdhs are candidates for determining neuronal molecular diversity. Here, we showed that the targeted deletion of DNase I hypersensitive (HS) site HS5-1, previously identified as a Pcdh-α regulatory element in vitro, affects especially the expression of specific Pcdh-α isoforms in vivo. We also identified a Pcdh-β cluster control region (CCR) containing six HS sites (HS16, 17, 17', 18, 19, and 20) downstream of the Pcdh-γ cluster. This CCR comprehensively activates the expression of the Pcdh-β gene cluster in cis, and its deletion dramatically decreases their expression levels. Deleting the CCR nonuniformly down-regulates some Pcdh-γ isoforms and does not affect Pcdh-α expression. Thus, the CCR effect extends beyond the 320-kb region containing the Pcdh-γ cluster to activate the upstream Pcdh-β genes. Thus, we concluded that the CCR is a highly specific regulatory unit for Pcdh-β expression on the clustered Pcdh genomic locus. These findings suggest that each Pcdh cluster is controlled by distinct regulatory elements that activate their expression and that the stochastic gene regulation of the clustered Pcdhs is controlled by the complex chromatin architecture of the clustered Pcdh locus.

A role for endosomal proteins in alphavirus dissemination in mosquitoes

Little is known about endosomal pathway proteins involved in arthropod-borne virus (arbovirus) assembly and cell-to-cell spread in vector mosquitoes. UNC93A and synaptic vesicle-2 (SV2) proteins are involved in intracellular transport in mammals. They show amino acid sequence conservation from mosquitoes to humans, and their transcripts are highly enriched in Aedes aegypti during arbovirus infection. Transient gene silencing of SV2 or UNC93A in mosquitoes infected with the recombinant alphavirus Sindbis MRE16-enhanced green fluorescent protein (SINV; family Togaviridae) resulted in the accumulation of viral positive- and negative-strand RNA, congregation of virus envelope antigen in intracellular networks, and reduced virus dissemination outside of the midgut. Further, UNC93A silencing, but not SV2 silencing, resulted in a 10-fold reduction in viral titres at 4 days post-infection. Together, these data support a role for UNC93A and SV2 in virus assembly or budding. Cis-regulatory elements (CREs) were identified at the 5'-ends of genes from the original data set in which SV2 and UNC93A were identified. Common CREs at the 5'-end genomic regions of a subset of enriched transcripts support the hypothesis that UNC93A transcription may be co-regulated with that of other ion transport and endosomal trafficking proteins.

A regulon conserved in monocot and dicot plants defines a functional module in antifungal plant immunity

At least two components that modulate plant resistance against the fungal powdery mildew disease are ancient and have been conserved since the time of the monocot-dicot split (≈ 200 Mya). These components are the seven transmembrane domain containing MLO/MLO2 protein and the syntaxin ROR2/PEN1, which act antagonistically and have been identified in the monocot barley (Hordeum vulgare) and the dicot Arabidopsis thaliana, respectively. Additionally, syntaxin-interacting N-ethylmaleimide sensitive factor adaptor protein receptor proteins (VAMP721/722 and SNAP33/34) as well as a myrosinase (PEN2) and an ABC transporter (PEN3) contribute to antifungal resistance in both barley and/or Arabidopsis. Here, we show that these genetically defined defense components share a similar set of coexpressed genes in the two plant species, comprising a statistically significant overrepresentation of gene products involved in regulation of transcription, posttranslational modification, and signaling. Most of the coexpressed Arabidopsis genes possess a common cis-regulatory element that may dictate their coordinated expression. We exploited gene coexpression to uncover numerous components in Arabidopsis involved in antifungal defense. Together, our data provide evidence for an evolutionarily conserved regulon composed of core components and clade/species-specific innovations that functions as a module in plant innate immunity.

Isolation, Characterization, and Expression Analysis of an NAP-Like cDNA from Crocus (Crocus sativus L.)

In Arabidopsis, it has been shown that the B-class MADS-box genes are expressed in the developing whorl 2 petals and whorl 3 stamens throughout the ontogeny of these organs. APETALA3/PISTILLATA (AP3/PI) heterodimers act as an inducer of a regulator-coding gene called NAP, a homologue to NAC-like transcription factor genes, required for meristem establishment, separation of floral organs, and leaf senescence. In monocots like crocus (Crocus sativus) cultivated for saffron production, the expression of B-class MADS-box CsatAP3/CsatPI genes extends to whorl 1 where petaloid tepals are formed. We report here for the first time the cloning and characterization of an NAC-like gene, named CsatNAP, from crocus. The sequence alignment indicated that CsatNAP protein contains the typical domain structure of plant NAC proteins consisted of the conserved five subdomains of the N-terminal NAC domain. Phylogenetic analysis revealed that CsatNAP protein falls in the subgroup NAP of the NAC proteins. Expression analysis of crocus NAP indicates that it is expressed in whorl 1, supporting the hypothesis that also in crocus, AP3/PI-like is associated with the expression of Crocus NAP gene. The expression patterns of CsatNAP cDNA were studied in flowers and different flower organs as well as in leaves and different part of leaves during senescence. The CsatNAP transcripts were detected in whole flowers and flower organs. Promoter analysis of CsatNAP revealed a number of putative common cis-regulatory elements and, among them, two CArG boxes indicative of its control by MADS box transcription factors known to bind on CArG sites. Micro-RNA (miRNA) target analysis showed that the NAP sequence of Crocus contain a possible target site for miRNA164. Furthermore, CsatNAP showed increased expression in senescence leaves compared to the green ones.

Application of Biostatistics and Bioinformatics Tools to Identify Putative Transcription Factor-Gene Regulatory Network of Ankylosing Spondylitis and Sarcoidosis

Transcription factors and corresponding cis-regulatory elements are considered key components in gene regulation. We combined biostatistics and bioinformatics tools to streamline identification of putative transcription factor-gene regulatory networks unique for two immune-mediated diseases, ankylosing spondylitis and sarcoidosis. After identifying differentially expressed genes from microarrays, we employed tightCluster to find tight clusters of potentially co-regulated genes. By subsequently applying bioinformatics tools to search for common cis-regulatory elements, putative transcription factor-gene regulatory networks were found. Recognition of these networks by applying this methodology could pave the way for new insights into disease pathogenesis.