BaRTv1.0:an improved barley reference transcript dataset to determine accurate changes in the barley transcriptome using RNA-seq

Background: The time required to analyse RNA-seq data varies considerably, due to discrete steps for computational assembly, quantification of gene expression and splicing analysis. Recent fast non-alignment tools such as Kallisto and Salmon overcome these problems, but these tools require a high quality, comprehensive reference transcripts dataset (RTD), which are rarely available in plants.Results: A high-quality, non-redundant barley gene RTD and database (Barley Reference Transcripts - BaRTv1.0) has been generated. BaRTv1.0, was constructed from a range of tissues, cultivars and abiotic treatments and transcripts assembled and aligned to the barley cv. Morex reference genome (Mascher et al. Nature; 544: 427-433, 2017). Full-length cDNAs from the barley variety Haruna nijo (Matsumoto et al. Plant Physiol; 156: 20-28, 2011) determined transcript coverage, and high-resolution RT-PCR validated alternatively spliced (AS) transcripts of 86 genes in five different organs and tissue. These methods were used as benchmarks to select an optimal barley RTD. BaRTv1.0-Quantification of Alternatively Spliced Isoforms (QUASI) was also made to overcome inaccurate quantification due to variation in 5' and 3' UTR ends of transcripts. BaRTv1.0-QUASI was used for accurate transcript quantification of RNA-seq data of five barley organs/tissues. This analysis identified 20,972 significant differentially expressed genes, 2791 differentially alternatively spliced genes and 2768 transcripts with differential transcript usage.Conclusion: A high confidence barley reference transcript dataset consisting of 60,444 genes with 177,240 transcripts has been generated. Compared to current barley transcripts, BaRTv1.0 transcripts are generally longer, have less fragmentation and improved gene models that are well supported by splice junction reads. Precise transcript quantification using BaRTv1.0 allows routine analysis of gene expression and AS.</p

AMPK associates with and causes fragmentation of the Golgi by phosphorylating the guanine nucleotide exchange factor GBF1

AMP-activated protein kinase (AMPK) is an energy sensor that regulates cellular functions in response to changes in energy availability. However, whether AMPK activity is spatially regulated, and the implications for cell function, have been unclear. We now report that AMPK associates with the Golgi, and that its activation by two specific pharmacological activators leads to Golgi fragmentation similar to that caused by the antibiotic Golgicide A, an inhibitor of Golgi-specific Brefeldin A resistance factor-1 (GBF1), a guanine nucleotide exchange factor that targets ADP-ribosylation factor 1 (ARF1). Golgi fragmentation in response to AMPK activators is lost in cells carrying gene knockouts of AMPK-α subunits. AMPK has been previously reported to phosphorylate GBF1 at residue Thr1337, and its activation causes phosphorylation at that residue. Importantly, Golgi disassembly upon AMPK activation is blocked in cells expressing a non-phosphorylatable GBF1-T1337A mutant generated by gene editing. Furthermore, the trafficking of a plasma membrane-targeted protein through the Golgi complex is delayed by AMPK activation. Our findings provide a mechanism to link AMPK activation during cellular energy stress to downregulation of protein trafficking involving the Golgi

Genetic pathways controlling inflorescence architecture and development in wheat and barley

Modifications of inflorescence architecture have been crucial for the successful domestication of wheat and barley, which are central members of the Triticeae tribe that provide essential grains for the human diet. Investigation of the genes and alleles that underpin domestication-related traits has provided valuable insights into the molecular regulation of inflorescence development of the Triticeae, and further investigation of modified forms of architecture are proving to be equally fruitful. The identified genes are involved in diverse biological processes, including transcriptional regulation, hormone biosynthesis and metabolism, post-transcriptional and post-translational regulation, which alter inflorescence architecture by modifying the development and fertility of lateral organs, called spikelets and florets. Recent advances in sequencing capabilities and the generation of mutant populations are accelerating the identification of genes that influence inflorescence development, which is important given that genetic variation for this trait promises to be a valuable resource for optimizing grain production. This review assesses recent advances in our understanding of the genes controlling inflorescence development in wheat and barley, with the aim of highlighting the importance of improvements in developmental biology for optimizing the agronomic performance of staple crop plants.Adam Gauley and Scott A. Bode

Development of a highly potent and selective degrader of LRRK2

The discovery of disease-modifying therapies for Parkinson's Disease (PD) represents a critical need in neurodegenerative medicine. Genetic mutations in leucine-rich repeat kinase 2 (LRRK2) are risk factors for the development of PD, and some of these mutations have been linked to increased LRRK2 kinase activity and neuronal toxicity in cellular and animal models. Furthermore, LRRK2 function as a scaffolding protein in several pathways has been implicated as a plausible mechanism underlying neurodegeneration caused by LRRK2 mutations. Given that both the kinase activity and scaffolding function of LRRK2 have been linked to neurodegeneration, we developed proteolysis-targeting chimeras (PROTACs) targeting LRRK2. The degrader molecule JH-XII-03-02 (6) displayed high potency and remarkable selectivity for LRKK2 when assessed in a of 468 panel kinases and serves the dual purpose of eliminating both the kinase activity as well as the scaffolding function of LRRK2.</p

BaRTv1.0: an improved barley reference transcript dataset to determine accurate changes in the barley transcriptome using RNA-seq

AbstractBackgroundTime consuming computational assembly and quantification of gene expression and splicing analysis from RNA-seq data vary considerably. Recent fast non-alignment tools such as Kallisto and Salmon overcome these problems, but these tools require a high quality, comprehensive reference transcripts dataset (RTD), which are rarely available in plants.ResultsA high-quality, non-redundant barley gene RTD and database (Barley Reference Transcripts – BaRTv1.0) has been generated. BaRTv1.0, was constructed from a range of tissues, cultivars and abiotic treatments and transcripts assembled and aligned to the barley cv. Morex reference genome (Mascher et al., 2017). Full-length cDNAs from the barley variety Haruna nijo (Matsumoto et al., 2011) determined transcript coverage, and high-resolution RT-PCR validated alternatively spliced (AS) transcripts of 86 genes in five different organs and tissue. These methods were used as benchmarks to select an optimal barley RTD. BaRTv1.0-Quantification of Alternatively Spliced Isoforms (QUASI) was also made to overcome inaccurate quantification due to variation in 5’ and 3’ UTR ends of transcripts. BaRTv1.0-QUASI was used for accurate transcript quantification of RNA-seq data of five barley organs/tissues. This analysis identified 20,972 significant differentially expressed genes, 2,791 differentially alternatively spliced genes and 2,768 transcripts with differential transcript usage.ConclusionA high confidence barley reference transcript dataset consisting of 60,444 genes with 177,240 transcripts has been generated. Compared to current barley transcripts, BaRTv1.0 transcripts are generally longer, have less fragmentation and improved gene models that are well supported by splice junction reads. Precise transcript quantification using BaRTv1.0 allows routine analysis of gene expression and AS.</jats:sec

BaRTv1.0: an improved barley reference transcript dataset to determine accurate changes in the barley transcriptome using RNA-seq

Abstract Background The time required to analyse RNA-seq data varies considerably, due to discrete steps for computational assembly, quantification of gene expression and splicing analysis. Recent fast non-alignment tools such as Kallisto and Salmon overcome these problems, but these tools require a high quality, comprehensive reference transcripts dataset (RTD), which are rarely available in plants. Results A high-quality, non-redundant barley gene RTD and database (Barley Reference Transcripts – BaRTv1.0) has been generated. BaRTv1.0, was constructed from a range of tissues, cultivars and abiotic treatments and transcripts assembled and aligned to the barley cv. Morex reference genome (Mascher et al. Nature; 544: 427–433, 2017). Full-length cDNAs from the barley variety Haruna nijo (Matsumoto et al. Plant Physiol; 156: 20–28, 2011) determined transcript coverage, and high-resolution RT-PCR validated alternatively spliced (AS) transcripts of 86 genes in five different organs and tissue. These methods were used as benchmarks to select an optimal barley RTD. BaRTv1.0-Quantification of Alternatively Spliced Isoforms (QUASI) was also made to overcome inaccurate quantification due to variation in 5′ and 3′ UTR ends of transcripts. BaRTv1.0-QUASI was used for accurate transcript quantification of RNA-seq data of five barley organs/tissues. This analysis identified 20,972 significant differentially expressed genes, 2791 differentially alternatively spliced genes and 2768 transcripts with differential transcript usage. Conclusion A high confidence barley reference transcript dataset consisting of 60,444 genes with 177,240 transcripts has been generated. Compared to current barley transcripts, BaRTv1.0 transcripts are generally longer, have less fragmentation and improved gene models that are well supported by splice junction reads. Precise transcript quantification using BaRTv1.0 allows routine analysis of gene expression and AS. </jats:sec