TATA-Box Binding Protein O-GlcNAcylation at T114 Regulates Formation of the B-TFIID Complex and Is Critical for Metabolic Gene Regulation.

In eukaryotes, gene expression is performed by three RNA polymerases that are targeted to promoters by molecular complexes. A unique common factor, the TATA-box binding protein (TBP), is thought to serve as a platform to assemble pre-initiation complexes competent for transcription. Here, we describe a novel molecular mechanism of nutrient regulation of gene transcription by dynamic O-GlcNAcylation of TBP. We show that O-GlcNAcylation at T114 of TBP blocks its interaction with BTAF1, hence the formation of the B-TFIID complex, and its dynamic cycling on and off of DNA. Transcriptomic and metabolomic analyses of TBPT114A CRISPR/Cas9-edited cells showed that loss of O-GlcNAcylation at T114 increases TBP binding to BTAF1 and directly impacts expression of 408 genes. Lack of O-GlcNAcylation at T114 is associated with a striking reprogramming of cellular metabolism induced by a profound modification of the transcriptome, leading to gross alterations in lipid storage.

Sequence-Directed Action of RSC Remodeler and General Regulatory Factors Modulates +1 Nucleosome Position to Facilitate Transcription.

Accessible chromatin is important for RNA polymerase II recruitment and transcription initiation at eukaryotic promoters. We investigated the mechanistic links between promoter DNA sequence, nucleosome positioning, and transcription. Our results indicate that positioning of the transcription start site-associated +1 nucleosome in yeast is critical for efficient TBP binding and is driven by two key factors, the essential chromatin remodeler RSC and a small set of ubiquitous general regulatory factors (GRFs). Our findings indicate that the strength and directionality of RSC action on promoter nucleosomes depends on the arrangement and proximity of two specific DNA motifs. This, together with the effect on nucleosome position observed in double depletion experiments, suggests that, despite their widespread co-localization, RSC and GRFs predominantly act through independent signals to generate accessible chromatin. Our results provide mechanistic insight into how the promoter DNA sequence instructs trans-acting factors to control nucleosome architecture and stimulate transcription initiation.

SAGA Is a General Cofactor for RNA Polymerase II Transcription.

Prior studies suggested that SAGA and TFIID are alternative factors that promote RNA polymerase II transcription, with about 10% of genes in S. cerevisiae dependent on SAGA. We reassessed the role of SAGA by mapping its genome-wide location and role in global transcription in budding yeast. We find that SAGA maps to the UAS elements of most genes, overlapping with Mediator binding and irrespective of previous designations of SAGA- or TFIID-dominated genes. Disruption of SAGA through mutation or rapid subunit depletion reduces transcription from nearly all genes, measured by newly synthesized RNA. We also find that the acetyltransferase Gcn5 synergizes with Spt3 to promote global transcription and that Spt3 functions to stimulate TBP recruitment at all tested genes. Our data demonstrate that SAGA acts as a general cofactor required for essentially all RNA polymerase II transcription and is not consistent with the previous classification of SAGA- and TFIID-dominated genes.

TAF1 is needed for the proliferation and maturation of thyroid follicle cells via Notch signaling.

Thyroid dysgenesis (TD) is the common pathogenic mechanism of congenital hypothyroidism (CH). In addition, known pathogenic genes are limited to those that are directly involved in thyroid development. To identify additional candidate pathogenetic genes, we performed forward genetic screening for TD in zebrafish, followed by positional cloning. The candidate gene was confirmed in vitro using the Nthy-ori 3.1 cell line and in vivo using a zebrafish model organism. We obtained a novel zebrafish line with thyroid dysgenesis and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 (taf1) by positional cloning. Further molecular studies revealed that taf1 was needed for the proliferation of thyroid follicular cells by binding to the NOTCH1 promoter region. Knockdown of TAF1 impaired the proliferation and maturation of thyroid cells, thereby leading to thyroid dysplasia. This study showed that TAF1 promoted Notch signaling and that this association played a pivotal role in thyroid development.NEW & NOTEWORTHY In our study, we obtained a novel zebrafish line with thyroid dysgenesis (TD) and identified the candidate pathogenetic mutation TATA-box binding protein associated Factor 1 (taf1). Further researches revealed that taf1 was required for thyroid follicular cells by binding to the NOTCH1 promoter region. Our findings revealed a novel role of TAF1 in thyroid morphogenesis.

TBP facilitates RNA Polymerase I transcription following mitosis.

The TATA-box binding protein (TBP) is the sole transcription factor common in the initiation complexes of the three major eukaryotic RNA Polymerases (Pol I, II and III). Although TBP is central to transcription by the three RNA Pols in various species, the emergence of TBP paralogs throughout evolution has expanded the complexity in transcription initiation. Furthermore, recent studies have emerged that questioned the centrality of TBP in mammalian cells, particularly in Pol II transcription, but the role of TBP and its paralogs in Pol I transcription remains to be re-evaluated. In this report, we show that in murine embryonic stem cells TBP localizes onto Pol I promoters, whereas the TBP paralog TRF2 only weakly associates to the Spacer Promoter of rDNA, suggesting that it may not be able to replace TBP for Pol I transcription. Importantly, acute TBP depletion does not fully disrupt Pol I occupancy or activity on ribosomal RNA genes, but TBP binding in mitosis leads to efficient Pol I reactivation following cell division. These findings provide a more nuanced role for TBP in Pol I transcription in murine embryonic stem cells.

AtSNP_TATAdb: Candidate Molecular Markers of Plant Advantages Related to Single Nucleotide Polymorphisms within Proximal Promoters of Arabidopsis thaliana L.

The mainstream of the post-genome target-assisted breeding in crop plant species includes biofortification such as high-throughput phenotyping along with genome-based selection. Therefore, in this work, we used the Web-service Plant_SNP_TATA_Z-tester, which we have previously developed, to run a uniform in silico analysis of the transcriptional alterations of 54,013 protein-coding transcripts from 32,833 Arabidopsis thaliana L. genes caused by 871,707 SNPs located in the proximal promoter region. The analysis identified 54,993 SNPs as significantly decreasing or increasing gene expression through changes in TATA-binding protein affinity to the promoters. The existence of these SNPs in highly conserved proximal promoters may be explained as intraspecific diversity kept by the stabilizing natural selection. To support this, we hand-annotated papers on some of the Arabidopsis genes possessing these SNPs or on their orthologs in other plant species and demonstrated the effects of changes in these gene expressions on plant vital traits. We integrated in silico estimates of the TBP-promoter affinity in the AtSNP_TATAdb knowledge base and showed their significant correlations with independent in vivo experimental data. These correlations appeared to be robust to variations in statistical criteria, genomic environment of TATA box regions, plants species and growing conditions.

Calpains as novel players in the molecular pathogenesis of spinocerebellar ataxia type 17.

Spinocerebellar ataxia type 17 (SCA17) is a neurodegenerative disease caused by a polyglutamine-encoding trinucleotide repeat expansion in the gene of transcription factor TATA box-binding protein (TBP). While its underlying pathomechanism is elusive, polyglutamine-expanded TBP fragments of unknown origin mediate the mutant protein's toxicity. Calcium-dependent calpain proteases are protagonists in neurodegenerative disorders. Here, we demonstrate that calpains cleave TBP, and emerging C-terminal fragments mislocalize to the cytoplasm. SCA17 cell and rat models exhibited calpain overactivation, leading to excessive fragmentation and depletion of neuronal proteins in vivo. Transcriptome analysis of SCA17 cells revealed synaptogenesis and calcium signaling perturbations, indicating the potential cause of elevated calpain activity. Pharmacological or genetic calpain inhibition reduced TBP cleavage and aggregation, consequently improving cell viability. Our work underlines the general significance of calpains and their activating pathways in neurodegenerative disorders and presents these proteases as novel players in the molecular pathogenesis of SCA17.

Candidate SNP Markers Significantly Altering the Affinity of TATA-Binding Protein for the Promoters of Human Hub Genes for Atherogenesis, Atherosclerosis and Atheroprotection.

Atherosclerosis is a systemic disease in which focal lesions in arteries promote the build-up of lipoproteins and cholesterol they are transporting. The development of atheroma (atherogenesis) narrows blood vessels, reduces the blood supply and leads to cardiovascular diseases. According to the World Health Organization (WHO), cardiovascular diseases are the leading cause of death, which has been especially boosted since the COVID-19 pandemic. There is a variety of contributors to atherosclerosis, including lifestyle factors and genetic predisposition. Antioxidant diets and recreational exercises act as atheroprotectors and can retard atherogenesis. The search for molecular markers of atherogenesis and atheroprotection for predictive, preventive and personalized medicine appears to be the most promising direction for the study of atherosclerosis. In this work, we have analyzed 1068 human genes associated with atherogenesis, atherosclerosis and atheroprotection. The hub genes regulating these processes have been found to be the most ancient. In silico analysis of all 5112 SNPs in their promoters has revealed 330 candidate SNP markers, which statistically significantly change the affinity of the TATA-binding protein (TBP) for these promoters. These molecular markers have made us confident that natural selection acts against underexpression of the hub genes for atherogenesis, atherosclerosis and atheroprotection. At the same time, upregulation of the one for atheroprotection promotes human health.

Core Promoter Regions of Antisense and Long Intergenic Non-Coding RNAs.

RNA polymerase II (POL II) is responsible for the transcription of messenger RNAs (mRNAs) and long non-coding RNAs (lncRNAs). Previously, we have shown the evolutionary invariance of the structural features of DNA in the POL II core promoters of the precursors of mRNAs. In this work, we have analyzed the POL II core promoters of the precursors of lncRNAs in Homo sapiens and Mus musculus genomes. Structural analysis of nucleotide sequences in positions -50, +30 bp in relation to the TSS have shown the extremely heterogeneous 3D structure that includes two singular regions - hexanucleotide "INR" around the TSS and octanucleotide "TATA-box" at around ~-28 bp upstream. Thus, the 3D structure of core promoters of lncRNA resembles the architecture of the core promoters of mRNAs; however, textual analysis revealed differences between promoters of lncRNAs and promoters of mRNAs, which lies in their textual characteristics; namely, the informational entropy at each position of the nucleotide text of lncRNA core promoters (by the exception of singular regions) is significantly higher than that of the mRNA core promoters. Another distinguishing feature of lncRNA is the extremely rare occurrence in the TATA box of octanucleotides with the consensus sequence. These textual differences can significantly affect the efficiency of the transcription of lncRNAs.

Dysfunction of CYC2g is responsible for the evolutionary shift from radiate to disciform flowerheads in the Chrysanthemum group (Asteraceae: Anthemideae).

Evolutionary shifts among radiate, disciform and discoid flowerheads have occurred repeatedly in a number of major lineages across the Asteraceae phylogeny; such transitions may also appear within evolutionarily young groups. Although several studies have demonstrated that CYC2 genes partake in regulating floral morphogenesis in Asteraceae, the evolution of capitulum forms within a recently diverging lineage has remained poorly understood. Here, we study the molecular regulation of the shift from a radiate to a disciform capitulum within the Chrysanthemum group. This is a recently radiating group mainly comprising two genera, Chrysanthemum and Ajania, that are phylogenetically intermingled but distinct in flowerhead morphology: Chrysanthemum spp. with radiate capitula and Ajania spp. with disciform capitula. We found that the morphogenesis of zygomorphy in the marginal floret in Ajania was disrupted soon after floral primordium emergence; CYC2g, one of the CYC2 copies that was expressed prominently in the ray floret of Chrysanthemum was not expressed in flowerheads of Ajania. Weakening the expression of ClCYC2g in Chrysanthemum lavandulifolium led to the gradual transition of a ray flower toward the disc-like form. Molecular evolutionary analyses indicated that the disciform capitulum might have evolved only once, approximately 8 Mya, arising from dysfunction of the CYC2g orthologs. A 20-nt deletion, including a putative TATA-box of the Ajania-type CYC2g promoter, appeared to inhibit the expression of the gene. Considering the divergent habitats of Chrysanthemum and Ajania, we propose that the shift from radiate to disciform capitulum must have been related to changes in pollination strategies under selective pressure.

iPromoter-Seqvec: identifying promoters using bidirectional long short-term memory and sequence-embedded features.

BACKGROUND: Promoters, non-coding DNA sequences located at upstream regions of the transcription start site of genes/gene clusters, are essential regulatory elements for the initiation and regulation of transcriptional processes. Furthermore, identifying promoters in DNA sequences and genomes significantly contributes to discovering entire structures of genes of interest. Therefore, exploration of promoter regions is one of the most imperative topics in molecular genetics and biology. Besides experimental techniques, computational methods have been developed to predict promoters. In this study, we propose iPromoter-Seqvec - an efficient computational model to predict TATA and non-TATA promoters in human and mouse genomes using bidirectional long short-term memory neural networks in combination with sequence-embedded features extracted from input sequences. The promoter and non-promoter sequences were retrieved from the Eukaryotic Promoter database and then were refined to create four benchmark datasets. RESULTS: The area under the receiver operating characteristic curve (AUCROC) and the area under the precision-recall curve (AUCPR) were used as two key metrics to evaluate model performance. Results on independent test sets showed that iPromoter-Seqvec outperformed other state-of-the-art methods with AUCROC values ranging from 0.85 to 0.99 and AUCPR values ranging from 0.86 to 0.99. Models predicting TATA promoters in both species had slightly higher predictive power compared to those predicting non-TATA promoters. With a novel idea of constructing artificial non-promoter sequences based on promoter sequences, our models were able to learn highly specific characteristics discriminating promoters from non-promoters to improve predictive efficiency. CONCLUSIONS: iPromoter-Seqvec is a stable and robust model for predicting both TATA and non-TATA promoters in human and mouse genomes. Our proposed method was also deployed as an online web server with a user-friendly interface to support research communities. Links to our source codes and web server are available at https://github.com/mldlproject/2022-iPromoter-Seqvec .

Karyopherin Kap114p-mediated trans-repression controls ribosomal gene expression under saline stress.

Nuclear accessibility of transcription factors controls gene expression, co-regulated by Ran-dependent nuclear localization and a competitive regulatory network. Here, we reveal that nuclear import factor-facilitated transcriptional repression attenuates ribosome biogenesis under chronic salt stress. Kap114p, one of the karyopherin-ßs (Kap-ßs) that mediates nuclear import of yeast TATA-binding protein (yTBP), exhibits a yTBP-binding affinity four orders of magnitude greater than its counterparts and suppresses binding of yTBP with DNA. Our crystal structure of Kap114p reveals an extensively negatively charged concave surface, accounting for high-affinity basic-protein binding. KAP114 knockout in yeast leads to a high-salt growth defect, with transcriptomic analyses revealing that Kap114p modulates expression of genes associated with ribosomal biogenesis by suppressing yTBP binding to target promoters, a trans-repression mechanism we attribute to reduced nuclear Ran levels under salinity stress. Our findings reveal that Ran integrates the nuclear transport pathway and transcription regulatory network, allowing yeast to respond to environmental stresses.

Genome-Wide Prediction of Transcription Start Sites in Conifers.

The identification of promoters is an essential step in the genome annotation process, providing a framework for gene regulatory networks and their role in transcription regulation. Despite considerable advances in the high-throughput determination of transcription start sites (TSSs) and transcription factor binding sites (TFBSs), experimental methods are still time-consuming and expensive. Instead, several computational approaches have been developed to provide fast and reliable means for predicting the location of TSSs and regulatory motifs on a genome-wide scale. Numerous studies have been carried out on the regulatory elements of mammalian genomes, but plant promoters, especially in gymnosperms, have been left out of the limelight and, therefore, have been poorly investigated. The aim of this study was to enhance and expand the existing genome annotations using computational approaches for genome-wide prediction of TSSs in the four conifer species: loblolly pine, white spruce, Norway spruce, and Siberian larch. Our pipeline will be useful for TSS predictions in other genomes, especially for draft assemblies, where reliable TSS predictions are not usually available. We also explored some of the features of the nucleotide composition of the predicted promoters and compared the GC properties of conifer genes with model monocot and dicot plants. Here, we demonstrate that even incomplete genome assemblies and partial annotations can be a reliable starting point for TSS annotation. The results of the TSS prediction in four conifer species have been deposited in the Persephone genome browser, which allows smooth visualization and is optimized for large data sets. This work provides the initial basis for future experimental validation and the study of the regulatory regions to understand gene regulation in gymnosperms.

OsTBP2.1, a TATA-Binding Protein, Alters the Ratio of OsNRT2.3b to OsNRT2.3a and Improves Rice Grain Yield.

The OsNRT2.3a and OsNRT2.3b isoforms play important roles in the uptake and transport of nitrate during rice growth. However, it is unclear which cis-acting element controls the transcription of OsNRT2.3 into these specific isoforms. In this study, we used a yeast one-hybrid assay to obtain the TATA-box binding protein OsTBP2.1, which binds to the TATA-box of OsNRT2.3, and verified its important role through transient expression and RNA-seq. We found that the TATA-box of OsNRT2.3 mutants and binding protein OsTBP2.1 together increased the transcription ratio of OsNRT2.3b to OsNRT2.3a. The overexpression of OsTBP2.1 promoted nitrogen uptake and increased rice yield compared with the wild-type; however, the OsTBP2.1 T-DNA mutant lines exhibited the opposite trend. Detailed analyses demonstrated that the TATA-box was the key cis-regulatory element for OsNRT2.3 to be transcribed into OsNRT2.3a and OsNRT2.3b. Additionally, this key cis-regulatory element, together with the binding protein OsTBP2.1, promoted the development of rice and increased grain yield.

Plant_SNP_TATA_Z-Tester: A Web Service That Unequivocally Estimates the Impact of Proximal Promoter Mutations on Plant Gene Expression.

Synthetic targeted optimization of plant promoters is becoming a part of progress in mainstream postgenomic agriculture along with hybridization of cultivated plants with wild congeners, as well as marker-assisted breeding. Therefore, here, for the first time, we compiled all the experimental data-on mutational effects in plant proximal promoters on gene expression-that we could find in PubMed. Some of these datasets cast doubt on both the existence and the uniqueness of the sought solution, which could unequivocally estimate effects of proximal promoter mutation on gene expression when plants are grown under various environmental conditions during their development. This means that the inverse problem under study is ill-posed. Furthermore, we found experimental data on in vitro interchangeability of plant and human TATA-binding proteins allowing the application of Tikhonov's regularization, making this problem well-posed. Within these frameworks, we created our Web service Plant_SNP_TATA_Z-tester and then determined the limits of its applicability using those data that cast doubt on both the existence and the uniqueness of the sought solution. We confirmed that the effects (of proximal promoter mutations on gene expression) predicted by Plant_SNP_TATA_Z-tester correlate statistically significantly with all the experimental data under study. Lastly, we exemplified an application of Plant_SNP_TATA_Z-tester to agriculturally valuable mutations in plant promoters.

A homozygous variant in TBPL2 was identified in women with oocyte maturation defects and infertility.

STUDY QUESTION: What are the genetic causes of oocyte maturation defects? SUMMARY ANSWER: A homozygous splicing variant (c.788 + 3A>G) in TATA-box binding protein like 2 (TBPL2) was identified as a contributory genetic factor in oocyte maturation defects. WHAT IS KNOWN ALREADY: TBPL2, a vertebrate oocyte-specific general transcription factor, is essential for oocyte development. TBPL2 variants have not been studied in human oocyte maturation defects. STUDY DESIGN, SIZE, DURATION: Two infertile families characterized by oocyte maturation defects were recruited for whole-exome sequencing (WES). PARTICIPANTS/MATERIALS, SETTING, METHODS: Genomic DNA was extracted from peripheral blood for WES analysis. Sanger sequencing was performed for data validation. Pathogenicity of variants was predicted by in silico analysis. Minigene assay and single-oocyte RNA sequencing were performed to investigate the effects of the variant on mRNA integrity and oocyte transcriptome, respectively. MAIN RESULTS AND THE ROLE OF CHANCE: A homozygous splicing variant (c.788 + 3A>G) in TBPL2 was identified in two unrelated families characterized by oocyte maturation defects. Haplotype analysis indicated that the disease allele of Families 1 and 2 was independent. The variant disrupted the integrity of TBPL2 mRNA. Transcriptome sequencing of affected oocytes showed that vital genes for oocyte maturation and fertilization were widely and markedly downregulated, suggesting that a mutation in the transcriptional factor, TBPL2, led to global gene alterations in oocytes. LIMITATIONS, REASONS FOR CAUTION: Limitations include the lack of direct functional evidence. Owing to the scarcity of human oocyte samples, only two immature MI oocytes were obtained from the patients, and we could only investigate the effect of the mutation at the transcriptional level by high-throughput sequencing technology. No extra oocytes were obtained to assess the transcriptional activity of the mutant oocytes by immunofluorescence, or investigate the effects on the binding of TBPL2 caused by the mutation. WIDER IMPLICATIONS OF THE FINDINGS: Our findings highlight a critical role of TBPL2 in female reproduction and identify a homozygous splicing mutation in TBPL2 that might be related to defects in human oocyte maturation. This information will facilitate the genetic diagnosis of infertile individuals with repeated failures of IVF, providing a basis for genetic counseling. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Key Research and Development Program of China (2018YFC1004000, 2017YFC1001504 and 2017YFC1001600), the National Natural Science Foundation of China (81871168, 31900409 and 31871509), the Foundation for Distinguished Young Scholars of Shandong Province (JQ201816), the Innovative Research Team of High-Level Local Universities in Shanghai (SSMU-ZLCX20180401) and the Fundamental Research Funds of Shandong University. The authors have no competing interests to declare. TRIAL REGISTRATION NUMBER: N/A.

Novel S. cerevisiae Hybrid Synthetic Promoters Based on Foreign Core Promoter Sequences.

Promoters are fundamental components of synthetic gene circuits. They are DNA segments where transcription initiation takes place. New constitutive and regulated promoters are constantly engineered in order to meet the requirements for protein and RNA expression into different genetic networks. In this work, we constructed and optimized new synthetic constitutive promoters for the yeast Saccharomyces cerevisiae. We started from foreign (e.g., viral) core promoters as templates. They are, usually, unfunctional in yeast but can be activated by extending them with a short sequence, from the CYC1 promoter, containing various transcription start sites (TSSs). Transcription was modulated by mutating the TATA box composition and varying its distance from the TSS. We found that gene expression is maximized when the TATA box has the form TATAAAA or TATATAA and lies between 30 and 70 nucleotides upstream of the TSS. Core promoters were turned into stronger promoters via the addition of a short UAS. In particular, the 40 nt bipartite UAS from the GPD promoter can enhance protein synthesis considerably when placed 150 nt upstream of the TATA box. Overall, we extended the pool of S. cerevisiae promoters with 59 new samples, the strongest overcoming the native TEF2 promoter.

Expression of the core promoter factors TATA box binding protein and TATA box binding protein-related factor 2 in Drosophila germ cells and their distinct functions in germline development.

In Drosophila, the expression of germline genes is initiated in primordial germ cells (PGCs) and is known to be associated with germline establishment. However, the transcriptional regulation of germline genes remains elusive. Previously, we found that the BTB/POZ-Zn-finger protein, Mamo, is necessary for the expression of the germline gene, vasa, in PGCs. Moreover, truncated Mamo lacking the BTB/POZ domain (MamoAF) is a potent vasa activator. In this study, we investigated the genetic interaction between MamoAF and specific transcriptional regulators to gain insight into the transcriptional regulation of germline development. We identified a general transcription factor, TATA box binding protein (TBP)-associated factor 3 (TAF3/BIP2), and a member of the TBP-like proteins, TBP-related factor 2 (TRF2), as new genetic modifiers of MamoAF. In contrast to TRF2, TBP was found to show no genetic interaction with MamoAF, suggesting that Trf2 has a selective function. Therefore, we focused on Trf2 expression and investigated its function in germ cells. We found that Trf2 mRNA, rather than Tbp mRNA, was preferentially expressed in PGCs during embryogenesis. Depletion of TRF2 in PGCs resulted in decreased mRNA expression of vasa. RNA interference-mediated knockdown showed that, while Trf2 is required for maintenance of germ cells, Tbp is needed for their differentiation during oogenesis. Therefore, these results suggest that Trf2 and Tbp expression is differentially regulated in germ cells and that these factors have distinct functions in Drosophila germline development.

Disruptive natural selection by male reproductive potential prevents underexpression of protein-coding genes on the human Y chromosome as a self-domestication syndrome.

BACKGROUND: In population ecology, the concept of reproductive potential denotes the most vital indicator of chances to produce and sustain a healthy descendant until his/her reproductive maturity under the best conditions. This concept links quality of life and longevity of an individual with disease susceptibilities encoded by his/her genome. Female reproductive potential has been investigated deeply, widely, and comprehensively in the past, but the male one has not received an equal amount of attention. Therefore, here we focused on the human Y chromosome and found candidate single-nucleotide polymorphism (SNP) markers of male reproductive potential. RESULTS: Examining in silico (i.e., using our earlier created Web-service SNP_TATA_Z-tester) all 1206 unannotated SNPs within 70 bp proximal promoters of all 63 Y-linked genes, we found 261 possible male-reproductive-potential SNP markers that can significantly alter the binding affinity of TATA-binding protein (TBP) for these promoters. Among them, there are candidate SNP markers of spermatogenesis disorders (e.g., rs1402972626), pediatric cancer (e.g., rs1483581212) as well as male anxiety damaging family relationships and mother's and children's health (e.g., rs187456378). First of all, we selectively verified in vitro both absolute and relative values of the analyzed TBP-promoter affinity, whose Pearson's coefficients of correlation between predicted and measured values were r = 0.84 (significance p <  0.025) and r = 0.98 (p <  0.025), respectively. Next, we found that there are twofold fewer candidate SNP markers decreasing TBP-promoter affinity relative to those increasing it, whereas in the genome-wide norm, SNP-induced damage to TBP-promoter complexes is fourfold more frequent than SNP-induced improvement (p <  0.05, binomial distribution). This means natural selection against underexpression of these genes. Meanwhile, the numbers of candidate SNP markers of an increase and decrease in male reproductive potential were indistinguishably equal to each other (p <  0.05) as if male self-domestication could have happened, with its experimentally known disruptive natural selection. Because there is still not enough scientific evidence that this could have happened, we discuss the human diseases associated with candidate SNP markers of male reproductive potential that may correspond to domestication-related disorders in pets. CONCLUSIONS: Overall, our findings seem to support a self-domestication syndrome with disruptive natural selection by male reproductive potential preventing Y-linked underexpression of a protein.

TATA-Like Boxes in RNA Polymerase III Promoters: Requirements for Nucleotide Sequences.

tRNA and some other non-coding RNA genes are transcribed by RNA polymerase III (pol III), due to the presence of intragenic promoter, consisting of boxes A and B spaced by 30-40 bp. Such pol III promoters, called type 2, are also intrinsic to Short Interspersed Elements (SINEs). The contribution of 5'-flanking sequences to the transcription efficiency of genes containing type 2 promoters is still studied insufficiently. Here, we studied this issue, focusing on the genes of two small non-coding RNAs (4.5SH and 4.5SI), as well as B1 and B2 SINEs from the mouse genome. We found that the regions from position -31 to -24 may significantly influence the transcription of genes and SINEs. We studied the influence of nucleotide substitutions in these sites, representing TATA-like boxes, on transcription of 4.5SH and 4.5SI RNA genes. As a rule, the substitutions of A and T to G or C reduced the transcription level, although the replacement of C with A also lowered it. In 4.5SH gene, five distal nucleotides of -31/-24 box (TTCAAGTA) appeared to be the most important, while in the box -31/-24 of 4.5SI gene (CTACATGA), all nucleotides, except for the first one, contributed significantly to the transcription efficiency. Random sequences occurring at positions -31/-24 upstream of SINE copies integrated into genome, promoted their transcription with different efficacy. In the 5'-flanking sequences of 4.5SH and 4.5SI RNA genes, the recognition sites of CREB, C/EBP, and Sp1 factors were found, and their deletion decreased the transcription.