The absence of the caffeine synthase gene is involved in the naturally decaffeinated status of Coffea humblotiana, a wild species from Comoro archipelago

Caffeine is the most consumed alkaloid stimulant in the world. It is synthesized through the activity of three known N-methyltransferase proteins. Here we are reporting on the 422-Mb chromosome-level assembly of the Coffea humblotiana genome, a wild and endangered, naturally caffeine-free, species from the Comoro archipelago. We predicted 32,874 genes and anchored 88.7% of the sequence onto the 11 chromosomes. Comparative analyses with the African Robusta coffee genome (C. canephora) revealed an extensive genome conservation, despite an estimated 11 million years of divergence and a broad diversity of genome sizes within the Coffea genus. In this genome, the absence of caffeine is likely due to the absence of the caffeine synthase gene which converts theobromine into caffeine through an illegitimate recombination mechanism. These findings pave the way for further characterization of caffeine-free species in the Coffea genus and will guide research towards naturally-decaffeinated coffee drinks for consumers.

Four large indels detected by cpTILLING in barley chloroplast mutator seedlings

In a previous work, a polymorphism detection strategy based on mismatch digestion was applied to the chloroplast genome of barley seedlings that carried the chloroplast mutator (cpm) genotype through many generations. Sixty-two different one- or two-nucleotide-polymorphisms were detected along with four large indels: an insertion of 15 bp in the intergenic region between tRNAHis and rps19 genes, a deletion of 620 bp in the psbA gene, a deletion of 79 bp in the intergenic region between rpl33 and rps18 genes and a deletion of 45 bp in the rps3 gene. In the present investigation, we analyzed direct repeats located at the borders of those four large indels. Furthermore, we investigated the consequences of protein expression of large indels located in coding regions. The deletion of 620 bp in the psbA gene was lethal at the second leaf stage when homoplastomic. The deletion of 45 bp in the rps3 gene, which eliminates 15 amino acids, did not affect the viability of the seedlings in homoplastomy. Interestingly, the deleted segment is also lacking in the wild type version of the rps3 gene of maize and sorghum. The presence of direct repeats at the borders of the four large indels suggests that they could have originated by illegitimate recombination. This would be in agreement with a previous hypothesis that the Cpm gene product would correspond to a mismatch repair (MMR) protein devoted to maintain plastome stability by playing fundamental roles in mismatch repair during replication and avoiding illegitimate recombination.

Understanding Rapid Transmitting, Invasive, Corona Pandemics through (Aair), (IS::Tn::IS), Crispr/Cas-9 DNA Editing

Antiadherent Immune Response (AAIR) against serotype 026: EPEC (Enteropathogenic (invasive) Escherichia coli) a fatal diarrhoea causing E.coli was successful in Balb/c mice experiment by the author. IS(Insertion Sequence) represented by IS1, IS2, IS3,-..IS10 flanking transposons (Tn) “IS::Tn::IS” and its illegitimate recombination was also studied curiously by the author to observe their spontaneous jumping, illegitimate recombination activities among DNA, chromosome and plasmids in Escherichia coli. Considering the fatality rate of corona as pandemic globally, the author has attempted to realise the possible application of AAIR, IS::Tn::IS and Crispr/Cas-9 in designing vaccine against corona, to prevent corona virus not to adhere in trachea and lung cells, to repair and to bring healthy life to mankind. is-Tn-is was observed/studied/discovered in Maize by Barbara Mc Clintock, USA, Peter Starlinger and Heinz Saedler, Germany studied the same in bacteria. Due to spontaneous mutations in corona, it is speculated by the author, that “IS::Tn::IS” DNA sequences might be present in corona, to mutate and to change the adhering, invasive and infective spike protein. The presence of IS-Tn-IS sequence have not been studied in SARS-COV-2, COVID-19, and corona. The sequence length, varied from IS1 (0.8kb) to IS10, (1.2 kb) (kilo base pairs) showed the potentiality spontaneous to do illegitimate recombination. If any of the sequence IS1, IS2, IS3…IS10 found common among HIV, MERS, SARS, Influenza, and H1N1 viruses, it could be useful to develop AAIR. Both AAIR vaccine concept and recent revolution in DNA editing, Crispr/cas-9(Clustered, Repeat Interspaced Short Palindromic Repeat), / (Crispr-associated protein-9) both could essentially be used, bidirectional in corona vaccine to protect pandemics and their repeats in future. The IR mechanisms which is proposed, that IS::Tn::IS cloned spike pathogenic DNA as involved in AAIR monoclonal antibody will block the adherence of corona spikes and Crispr/cas-9 will repair, the transmitted, invasive corona, in human body by programmable DNA of non-pathogenic influenza virus. The author describes few of those possible mechanisms of Is:: spike protein and Crispr based transposons to mobilize IR vaccine against corona, considering the nanostructures of corona and the use of it huge surface area in immune surveillance.

RAP80 and BRCA1 PARsylation protect chromosome integrity by preventing retention of BRCA1-B/C complexes in DNA repair foci

BRCA1 promotes error-free, homologous recombination-mediated repair (HRR) of DNA double-stranded breaks (DSBs). When excessive and uncontrolled, BRCA1 HRR activity promotes illegitimate recombination and genome disorder. We and others have observed that the BRCA1-associated protein RAP80 recruits BRCA1 to postdamage nuclear foci, and these chromatin structures then restrict the amplitude of BRCA1-driven HRR. What remains unclear is how this process is regulated. Here we report that both BRCA1 poly-ADP ribosylation (PARsylation) and the presence of BRCA1-bound RAP80 are critical for the normal interaction of BRCA1 with some of its partners (e.g., CtIP and BACH1) that are also known components of the aforementioned focal structures. Surprisingly, the simultaneous loss of RAP80 and failure therein of BRCA1 PARsylation results in the dysregulated accumulation in these foci of BRCA1 complexes. This in turn is associated with the intracellular development of a state of hyper-recombination and gross chromosomal disorder. Thus, physiological RAP80-BRCA1 complex formation and BRCA1 PARsylation contribute to the kinetics by which BRCA1 HRR-sustaining complexes normally concentrate in nuclear foci. These events likely contribute to aneuploidy suppression.

Genome comparison implies the role of Wsm2 in membrane trafficking and protein degradation

Wheat streak mosaic virus (WSMV) causes streak mosaic disease in wheat (Triticum aestivum L.) and has been an important constraint limiting wheat production in many regions around the world. Wsm2 is the only resistance gene discovered in wheat genome and has been located in a short genomic region of its chromosome 3B. However, the sequence nature and the biological function of Wsm2 remain unknown due to the difficulty of genetic manipulation in wheat. In this study, we tested WSMV infectivity among wheat and its two closely related grass species, rice (Oryza sativa) and Brachypodium distachyon. Based on the phenotypic result and previous genomic studies, we developed a novel bioinformatics pipeline for interpreting a potential biological function of Wsm2 and its ancestor locus in wheat. In the WSMV resistance tests, we found that rice has a WMSV resistance gene while Brachypodium does not, which allowed us to hypothesize the presence of a Wsm2 ortholog in rice. Our OrthoMCL analysis of protein coding genes on wheat chromosome 3B and its syntenic chromosomes in rice and Brachypodium discovered 4,035 OrthoMCL groups as preliminary candidates of Wsm2 orthologs. Given that Wsm2 is likely duplicated through an intrachromosomal illegitimate recombination and that Wsm2 is dominant, we inferred that this new WSMV-resistance gene acquired an activation domain, lost an inhibition domain, or gained high expression compared to its ancestor locus. Through comparison, we identified that 67, 16, and 10 out of 4,035 OrthoMCL orthologous groups contain a rice member with 25% shorter or longer in length, or 10 fold more expression, respectively, than those from wheat and Brachypodium. Taken together, we predicted a total of 93 good candidates for a Wsm2 ancestor locus. All of these 93 candidates are not tightly linked with Wsm2, indicative of the role of illegitimate recombination in the birth of Wsm2. Further sequence analysis suggests that the protein products of Wsm2 may combat WSMV disease through a molecular mechanism involving protein degradation and/or membrane trafficking. The 93 putative Wsm2 ancestor loci discovered in this study could serve as good candidates for future genetic isolation of the true Wsm2 locus.

Genome comparison implies the role of Wsm2 in membrane trafficking and protein degradation

Wheat streak mosaic virus (WSMV) causes streak mosaic disease in wheat (Triticum aestivum L.) and has been an important constraint limiting wheat production in many regions around the world. Wsm2 is the only resistance gene discovered in wheat genome and has been located in a short genomic region of its chromosome 3B. However, the sequence nature and the biological function of Wsm2 remain unknown due to the difficulty of genetic manipulation in wheat. In this study, we tested WSMV infectivity among wheat and its two closely related grass species, rice (Oryza sativa) and Brachypodium distachyon. Based on the phenotypic result and previous genomic studies, we developed a novel bioinformatics pipeline for interpreting a potential biological function of Wsm2 and its ancestor locus in wheat. In the WSMV resistance tests, we found that rice has a WMSV resistance gene while Brachypodium does not, which allowed us to hypothesize the presence of a Wsm2 ortholog in rice. Our OrthoMCL analysis of protein coding genes on wheat chromosome 3B and its syntenic chromosomes in rice and Brachypodium discovered 4,035 OrthoMCL groups as preliminary candidates of Wsm2 orthologs. Given that Wsm2 is likely duplicated through an intrachromosomal illegitimate recombination and that Wsm2 is dominant, we inferred that this new WSMV-resistance gene acquired an activation domain, lost an inhibition domain, or gained high expression compared to its ancestor locus. Through comparison, we identified that 67, 16, and 10 out of 4,035 OrthoMCL orthologous groups contain a rice member with 25% shorter or longer in length, or 10 fold more expression, respectively, than those from wheat and Brachypodium. Taken together, we predicted a total of 93 good candidates for a Wsm2 ancestor locus. All of these 93 candidates are not tightly linked with Wsm2, indicative of the role of illegitimate recombination in the birth of Wsm2. Further sequence analysis suggests that the protein products of Wsm2 may combat WSMV disease through a molecular mechanism involving protein degradation and/or membrane trafficking. The 93 putative Wsm2 ancestor loci discovered in this study could serve as good candidates for future genetic isolation of the true Wsm2 locus.