The Landscape of Repetitive Elements in the Refined Genome of Chilli Anthracnose Fungus Colletotrichum truncatum

The ascomycete fungus Colletotrichum truncatum is a major phytopathogen with a broad host range which causes anthracnose disease of chilli. The genome sequencing of this fungus led to the discovery of functional categories of genes that may play important roles in fungal pathogenicity. However, the presence of gaps in C. truncatum draft assembly prevented the accurate prediction of repetitive elements, which are the key players to determine the genome architecture and drive evolution and host adaptation. We re-sequenced its genome using single-molecule real-time (SMRT) sequencing technology to obtain a refined assembly with lesser and smaller gaps and ambiguities. This enabled us to study its genome architecture by characterising the repetitive sequences like transposable elements (TEs) and simple sequence repeats (SSRs), which constituted 4.9 and 0.38% of the assembled genome, respectively. The comparative analysis among different Colletotrichum species revealed the extensive repeat rich regions, dominated by Gypsy superfamily of long terminal repeats (LTRs), and the differential composition of SSRs in their genomes. Our study revealed a recent burst of LTR amplification in C. truncatum, C. higginsianum, and C. scovillei. TEs in C. truncatum were significantly associated with secretome, effectors and genes in secondary metabolism clusters. Some of the TE families in C. truncatum showed cytosine to thymine transitions indicative of repeat-induced point mutation (RIP). C. orbiculare and C. graminicola showed strong signatures of RIP across their genomes and “two-speed” genomes with extensive AT-rich and gene-sparse regions. Comparative genomic analyses of Colletotrichum species provided an insight into the species-specific SSR profiles. The SSRs in the coding and non-coding regions of the genome revealed the composition of trinucleotide repeat motifs in exons with potential to alter the translated protein structure through amino acid repeats. This is the first genome-wide study of TEs and SSRs in C. truncatum and their comparative analysis with six other Colletotrichum species, which would serve as a useful resource for future research to get insights into the potential role of TEs in genome expansion and evolution of Colletotrichum fungi and for development of SSR-based molecular markers for population genomic studies

The nucleolar protein Esf2 interacts directly with the DExD/H box RNA helicase, Dbp8, to stimulate ATP hydrolysis

While 18 putative RNA helicases are involved in ribosome biogenesis in Saccharomyces cerevisiae, their enzymatic properties have remained largely biochemically uncharacterized. To better understand their function, we examined the enzymatic properties of Dpb8, a DExD/H box protein previously shown to be required for the synthesis of the 18S rRNA. As expected for an RNA helicase, we demonstrate that recombinant Dbp8 has ATPase activity in vitro, and that this activity is dependent on an intact ATPase domain. Strikingly, we identify Esf2, a nucleolar putative RNA binding protein, as a binding partner for Dbp8, and show that it enhances Dbp8 ATPase activity by decreasing the K(M) for ATP. Thus, we have uncovered Esf2 as the first example of a protein co-factor that has a stimulatory effect on a nucleolar RNA helicase. We show that Esf2 can bind to pre-rRNAs and speculate that it may function to bring Dbp8 to the pre-rRNA, thereby both regulating its enzymatic activity and guiding Dbp8 to its site of action

The nucleolar protein Esf2 interacts directly with the DExD/H box RNA helicase, Dbp8, to stimulate ATP hydrolysis

While 18 putative RNA helicases are involved in ribosome biogenesis in Saccharomyces cerevisiae, their enzymatic properties have remained largely biochemically uncharacterized. To better understand their function, we examined the enzymatic properties of Dpb8, a DExD/H box protein previously shown to be required for the synthesis of the 18S rRNA. As expected for an RNA helicase, we demonstrate that recombinant Dbp8 has ATPase activity in vitro, and that this activity is dependent on an intact ATPase domain. Strikingly, we identify Esf2, a nucleolar putative RNA binding protein, as a binding partner for Dbp8, and show that it enhances Dbp8 ATPase activity by decreasing the K(M) for ATP. Thus, we have uncovered Esf2 as the first example of a protein co-factor that has a stimulatory effect on a nucleolar RNA helicase. We show that Esf2 can bind to pre-rRNAs and speculate that it may function to bring Dbp8 to the pre-rRNA, thereby both regulating its enzymatic activity and guiding Dbp8 to its site of action

Denisova admixture and the first modern human dispersals into Southeast Asia and Oceania

It has recently been shown that ancestors of New Guineans and Bougainville Islanders have inherited a proportion of their ancestry from Denisovans, an archaic hominin group from Siberia. However, only a sparse sampling of populations from Southeast Asia and Oceania were analyzed. Here, we quantify Denisova admixture in 33 additional populations from Asia and Oceania. Aboriginal Australians, Near Oceanians, Polynesians, Fijians, east Indonesians, and Mamanwa (a ‘‘Negrito’’ group from the Philippines) have all inherited genetic material from Denisovans, but mainland East Asians, western Indonesians, Jehai (a Negrito group from Malaysia), and Onge (a Negrito group from the Andaman Islands) have not. These results indicate that Denisova gene flow occurred into the common ancestors of New Guineans, Australians, and Mamanwa but not into the ancestors of the Jehai and Onge and suggest that relatives of present-day East Asians were not in Southeast Asia when the Denisova gene flow occurred. Our finding that descendants of the earliest inhabitants of Southeast Asia do not all harbor Denisova admixture is inconsistent with a history in which the Denisova interbreeding occurred in mainland Asia and then spread over Southeast Asia, leading to all its earliest modern human inhabitants. Instead, the data can be most parsimoniously explained if the Denisova gene flow occurred in Southeast Asia itself. Thus, archaic Denisovans must have lived over an extraordinarily broad geographic and ecological range, from Siberia to tropical Asia

Evidence for an Arginine Exporter Encoded by yggA (argO) That Is Regulated by the LysR-Type Transcriptional Regulator ArgP in Escherichia coli

The anonymous open reading frame yggA of Escherichia coli was identified in this study as a gene that is under the transcriptional control of argP (previously called iciA), which encodes a LysR-type transcriptional regulator protein. Strains with null mutations in either yggA or argP were supersensitive to the arginine analog canavanine, and yggA-lac expression in vivo exhibited argP(+)-dependent induction by arginine. Lysine supplementation phenocopied the argP null mutation in that it virtually abolished yggA expression, even in the argP(+) strain. The dipeptides arginylalanine and lysylalanine behaved much like arginine and lysine, respectively, to induce and to turn off yggA transcription. Dominant missense mutations in argP (argP(d)) that conferred canavanine resistance and rendered yggA-lac expression constitutive were obtained. The protein deduced to be encoded by yggA shares similarity with a basic amino acid exporter (LysE) of Corynebacterium glutamicum, and we obtained evidence for increased arginine efflux from E. coli strains with either the argP(d) mutation or multicopy yggA(+). The null yggA mutation abolished the increased arginine efflux from the argP(d) strain. Our results suggest that yggA encodes an ArgP-regulated arginine exporter, and we have accordingly renamed it argO (for “arginine outward transport”). We propose that the physiological function of argO may be either to prevent the accumulation to toxic levels of canavanine (which is a plant-derived antimetabolite) or arginine or to maintain an appropriate balance between the intracellular lysine and arginine concentrations

Genome sequencing and comparative genomics reveal a repertoire of putative pathogenicity genes in chilli anthracnose fungus <i>Colletotrichum truncatum</i>

<div><p><i>Colletotrichum truncatum</i>, a major fungal phytopathogen, causes the anthracnose disease on an economically important spice crop chilli (<i>Capsicum annuum</i>), resulting in huge economic losses in tropical and sub-tropical countries. It follows a subcuticular intramural infection strategy on chilli with a short, asymptomatic, endophytic phase, which contrasts with the intracellular hemibiotrophic lifestyle adopted by most of the <i>Colletotrichum</i> species. However, little is known about the molecular determinants and the mechanism of pathogenicity in this fungus. A high quality whole genome sequence and gene annotation based on transcriptome data of an Indian isolate of <i>C</i>. <i>truncatum</i> from chilli has been obtained. Analysis of the genome sequence revealed a rich repertoire of pathogenicity genes in <i>C</i>. <i>truncatum</i> encoding secreted proteins, effectors, plant cell wall degrading enzymes, secondary metabolism associated proteins, with potential roles in the host-specific infection strategy, placing it next only to the <i>Fusarium</i> species. The size of genome assembly, number of predicted genes and some of the functional categories were similar to other sequenced <i>Colletotrichum</i> species. The comparative genomic analyses with other species and related fungi identified some unique genes and certain highly expanded gene families of CAZymes, proteases and secondary metabolism associated genes in the genome of <i>C</i>. <i>truncatum</i>. The draft genome assembly and functional annotation of potential pathogenicity genes of <i>C</i>. <i>truncatum</i> provide an important genomic resource for understanding the biology and lifestyle of this important phytopathogen and will pave the way for designing efficient disease control regimens.</p></div

Assembly statistics of the <i>C</i>. <i>truncatum</i> genome assembly.

<p>Assembly statistics of the <i>C</i>. <i>truncatum</i> genome assembly.</p

The synteny plot between <i>C</i>. <i>higginsianum</i> and <i>C</i>. <i>truncatum</i> genomes obtained using SyMAP.

<p>The circular plot shows some of the syntenic blocks between the scaffolds of <i>C</i>. <i>truncatum</i> in lower half of the circle mapping to the reference chromosomes of <i>C</i>. <i>higginsianum</i> in upper half joined with the lines of same colour as the reference.</p

A maximum likelihood tree of <i>Colletotrichum</i> species and other fungi with diverse lifestyles.

<p>Bootstrap support values (1000 replicates) of 100% were obtained at the nodes. <i>A</i>. <i>nidulans</i> was taken as an outgroup for the analysis. The <i>Colletotrichum</i> species complexes and the families are shown in parallel. <i>C</i>. <i>truncatum</i>, <i>C</i>. <i>gloeosporioides</i> and <i>C</i>. <i>orbiculare</i> form a separate clade in the <i>Colletotrichum</i> genus suggesting their origin from a common ancestor.</p