Biology subjects

Buchmann, G.

Publications and source records attributed to Buchmann, G..

10 recordsLinked to original sources

bioRxiv2026

A single-cell transcriptome atlas of the barley root apical meristem uncovers conserved and divergent roles of HvWOX5

Single-cell approaches have transformed plant developmental biology; however, cell-type-resolved resources for cereals remain limited. Here, we present a single-cell transcriptome atlas of the barley root apical meristem (RAM), which resolves 24 transcriptionally distinct cell populations. We assigned major root cell identities by integrating marker gene validation using Hybridization Chain Reaction (HCR) RNA fluorescence in situ hybridization and spatial transcriptomics with cross-species comparisons of published root atlases. Pseudotime analysis reconstructed developmental trajectories from the quiescent center to differentiating tissues, supporting the spatial and developmental organization of the atlas. We further demonstrated the utility of this resource by identifying HvWOX5 expression in the quiescent center and metaxylem and showing that HvWOX5 loss-of-function mutants displayed reduced root and meristem lengths, altered stem cell niche homeostasis, and disrupted metaxylem organization. Taken together, this atlas provides a framework for dissecting barley root development and identifies HvWOX5 as a key regulator of RAM organization and metaxylem patterning.

Plant Biology↗

bioRxiv2025

Golden Promise-rapid, a fast-cycling barley genotype with high transformation efficiency

The spring barley cultivar Golden Promise (GP) is the major reference genotype for transformation due to its high transformability and availability of a reference genome. However, GP is characterized by a long generation cycle and stress susceptibility under non-optimal growth conditions because it carries a mutation at the floral inducer Photoperiod-H1 (Ppd-H1). Previously, we showed that a GP introgression line, Golden Promise-fast (GP-fast), generated by introducing the wild-type Ppd-H1 allele from the winter barley cultivar Igri, exhibits early flowering and improved stress resilience. In this study, we generated a fast-cycling genotype, Golden Promise-rapid (GP-rapid), isogenic to GP with high transformation efficiency. We conducted two backcrosses of GP-fast to reduce the residual Igri genome. The resulting genotype contains only a single introgression of approximately 0.6 Mbp at the Ppd-H1 locus on chromosome 2H. Under speed breeding conditions, its generation time was reduced to 63 days (25% shorter than GPs 84 days). Parallel transformation of GP, GP-fast, and GP-rapid using CRISPR/Cas9-mediated genome editing of Ppd-H1 revealed high regeneration and transformation efficiencies of GP-rapid, comparable to GP. Overall, we report on the development of a fast-cycling GP isogenic line as a research tool for efficient generation of transgenic and gene-edited barley plants. HighlightsA new fast-cycling barley genotype, GP-rapid, reduces generation time by 25% while retaining high transformation efficiency, advancing functional genomic studies in barley.

plant biology↗

bioRxiv2024

From Flowering Time to High Temperature Resilience: PHOTOPERIOD 1 (PPD-H1) Controls Spike Fertility through Improving Stress Resistance and Energy Metabolism under High Ambient Temperatures

High ambient temperature (HT) impairs reproductive development and grain yield in temperate crops. To ensure reproductive success under HT, plants must maintain developmental stability. However, the mechanisms integrating plant development and temperature resilience are largely unknown. Here, we demonstrate that PHOTOPERIOD 1 (PPD-H1), homologous to PSEUDO RESPONSE REGULATOR genes of the Arabidopsis circadian clock, controls developmental stability in response to HT in barley. We analyzed HT responses in independent introgression lines with either the ancestral wild-type Ppd-H1 allele or the natural ppd-h1 variant, selected in spring varieties to delay flowering and enhance yield under favourable conditions. HT delayed inflorescence development and reduced grain number in ppd-h1 mutant lines, while the wild-type Ppd-H1 genotypes accelerated reproductive development and showed a stable grain set under HT. Using a CRISPR/Cas9-induced ppd-h1 mutant, we confirmed that the CCT domain of Ppd-H1 controls developmental stability, but not clock gene expression. Transcriptome and phytohormone analyses in developing leaves and inflorescences revealed increased stress gene expression and abscisic acid levels in the leaf and inflorescence of the natural and induced mutant ppd-h1 lines. Furthermore, the mutant ppd-h1 lines downregulated photosynthesis-and energy metabolism-related genes, and reduced auxin and cytokinin levels in the inflorescence, which impaired anther and pollen development. By contrast, in the wild-type Ppd-H1 plants, the transcriptome and phytohormone levels and anther and pollen development remained stable under HT. Our findings suggest that Ppd-H1 enhances stress resistance and energy metabolism, thereby stabilizing reproductive development, floret fertility and grain set under HT.

plant biology↗

bioRxiv2023

Deformed wing virus genotypes A and B do not elicit immunologically different responses in naive honey bee hosts

Deformed wing virus (DWV), in association with Varroa destructor, is currently the leading factor associated with global honey bee deaths. With the exception of Australia, the virus and mite have a near global distribution, making it difficult to separate the effect of one from the other. Over time, the prevalence of the two main DWV genotypes (DWV-A and DWV-B) has changed, leading to the suggestion that the two strains elicit a different immune response by the host, the western honey bee Apis mellifera. Here we use a honey bee population naive to both the mite and the virus to investigate if honey bees show a different immunological response to DWV genotypes. We examined the expression of 19 immune genes by RT-qPCR and comprehensively analysed the small RNA response in honey bees after experimental injection with DWV-A and DWV-B. We found no evidence to indicate that DWV-A and DWV-B elicit a different immune response in honey bees. We found that RNA interference genes are up-regulated during DWV infection and that the small interfering RNA (siRNA) response is proportional to viral loads, yet does not inhibit the virus from accumulating to high loads. We also found that the siRNA response towards DWV was weaker than the response to another honey bee pathogen, Black queen cell virus. This suggests that DWV is comparatively better at evading antiviral host defences. There was no evidence for the production of virus-derived PIWI-RNAs in response to DWV infection. In contrast to previous studies, and in the absence of V. destructor, we found no evidence that DWV has an immunosuppressive effect in honey bees. Overall, our results advance our understanding of the immunological effect DWV elicits in honey bees.

immunology↗

bioRxiv2023

Virus replication in the honey bee parasite, Varroa destructor

Arthropod vectors such as mites and ticks introduce an alternative viral transmission route between their hosts. The ectoparasitic mite Varroa destructor is the leading threat to the health of Western honey bees (Apis mellifera) primarily through its action as a vector of viruses. However, it is unclear whether viruses transmitted by V. destructor actively infect and replicate in mites, which could facilitate increased transmission and select for more virulent strains. To better understand the role of V. destructor as a vector, we took advantage of differences between bee and mite antiviral RNA interference pathways to identify the host specificity of replicating viruses. We used small RNA sequencing of individual V. destructor mites to examine viral small interfering RNA (vsiRNA) profiles of Deformed wing virus genotypes (DWV-A and DWV-B), associated with colony declines, as well as nine other viruses present in our samples. We found active replication of six V. destructor-associated viruses, including a novel virus, Varroa destructor virus 9 (VDV-9), and replication of two honey bee associated viruses, including both DWV-A and -B genotypes, suggesting that mites are biological vectors for important bee pathogens. We show that the antiviral RNAi response can be used define the host range of viruses in host-parasite interactions, such as honey bees and their parasites, enabling a better understanding of the role of a vector in the evolution and spread of honey bee pathogens.

genetics↗

bioRxiv2023

EARLY MATURITY 7 modifies the circadian clock and photoperiod sensitivity in barley

Breeding for variation in photoperiod response is crucial to adapt crop plants to novel environments. Plants measure changes in day length by the circadian clock, an endogenous timekeeper that allows plants to anticipate changes in diurnal and seasonal light-dark cycles. Here, we describe the early maturity 7 (eam7) mutation in barley, which interacts with natural variation at PHOTOPERIOD 1 (Ppd-H1) to cause early flowering independent of the photoperiod. We identify LIGHT-REGULATED WD 1 (LWD1) as a putative candidate to underly the eam7 locus in barley as supported by genetic mapping and CRISPR-Cas9 generated lwd1 mutants. Mutations in eam7 cause a significant phase advance and a misregulation of core clock and clock output genes under diurnal conditions. Early flowering correlated with an upregulation of Ppd-H1 during the night and consequent induction of the florigen FLOWERING LOCUS T1 under short days. We propose that EAM7 controls photoperiodic flowering in barley by controlling the light input into the clock and diurnal expression patterns of the major photoperiod response gene Ppd-H1.

plant biology↗

bioRxiv2022

Ageing impairs the neuro-vascular interface in the heart

Aging is a major risk factor for impaired cardiovascular health. The aging myocardium is characterized by electrophysiological dysfunctions such as a reduced heart rate variability. These alterations can be intrinsic within cardiomyocytes, but might be modulated by the cardiac autonomic nervous system, as well1. It is known that nerves align with vessels during development2, but the impact of aging on the cardiac neuro-vascular interface is unknown. Here, we report that aging reduces nerve density specifically in the left ventricle and dysregulates vascular-derived neuro-regulatory genes. Aging leads further to a down-regulation of miR-145 and de-repression of the neuro-repulsive factor Semaphorin-3A. miR-145 deletion increased Sema3a expression and reduced axon density, thus mimicking the observed aged heart phenotype. Removal of senescent cells, which accumulated with chronological age while nerve density declined, rescued from age-induced dennervation, reduced Sema3a expression and preserved heart rate variability. These data suggest that senescence-associated regulation of neuro-regulatory genes contributes to a declined nerve density of the aging heart and thereby to a reduced heart rate variability.

physiology↗

bioRxiv2022

The endothelial-specific LINC00607 mediates endothelial angiogenic function

Long non-coding RNAs (lncRNAs) can act as regulatory RNAs which, by altering the expression of target genes, impact on the cellular phenotype and cardiovascular disease development. Endothelial lncRNAs and their vascular functions are largely undefined. Deep RNA-Seq and FANTOM5 CAGE analysis revealed the lncRNA LINC00607 to be highly enriched in human endothelial cells. LINC00607 was induced in response to hypoxia, arteriosclerosis regression in non-human primates and also in response to propranolol used to induce regression of human arteriovenous malformations. siRNA knockdown or CRISPR/Cas9 knockout of LINC00607 attenuated VEGF-A-induced angiogenic sprouting. LINC00607 knockout in endothelial cells also integrated less into newly formed vascular networks in an in vivo assay in SCID mice. Overexpression of LINC00607 in CRISPR knockout cells restored normal endothelial function. RNA- and ATAC-Seq after LINC00607 knockout revealed changes in the transcription of endothelial gene sets linked to the endothelial phenotype and in chromatin accessibility around ERG-binding sites. Mechanistically, LINC00607 interacted with the SWI/SNF chromatin remodeling protein BRG1. CRISPR/Cas9-mediated knockout of BRG1 in HUVEC followed by CUT&RUN revealed that BRG1 is required to secure a stable chromatin state, mainly on ERG-binding sites. In conclusion, LINC00607 is an endothelial-enriched lncRNA that maintains ERG target gene transcription by interacting with the chromatin remodeler BRG1.

molecular biology↗

bioRxiv2021

Abundant small RNAs in the reproductive tissues of the honey bee, Apis mellifera, are a plausible mechanism for epigenetic inheritance and parental manipulation of gene expression

Polyandrous social insects such as the honey bee are prime candidates for parental manipulation of gene expression in offspring. Although there is good evidence for parent-of-origin effects in honey bees the epigenetic mechanisms that underlie these effects remain a mystery. Small RNA molecules such as miRNAs, piRNAs and siRNAs play important roles in transgenerational epigenetic inheritance and in the regulation of gene expression during development. Here we present the first characterisation of small RNAs present in honey bee reproductive tissues: ovaries, spermatheca, semen, fertilised and unfertilised eggs, and testes. We show that semen contains fewer piRNAs relative to eggs and ovaries, and that piRNAs and miRNAs which map antisense to genes involved in DNA regulation and developmental processes are differentially expressed between tissues. tRNA fragments are highly abundant in semen and have a similar profile to those seen in semen in other animals. Intriguingly we find abundant piRNAs that target the sex determination locus, suggesting that piRNAs may play a role in honey bee sex determination. We conclude that small RNAs play a fundamental role in honey bee gametogenesis and reproduction and provide a plausible mechanism for parent-of origin-effects on gene expression and reproductive physiology.

genomics↗

bioRxiv2018

The chromatin modifiers SET-25 and SET-32 are required for initiation but not long-term maintenance of transgenerational epigenetic inheritance

Some epigenetic modifications are inherited from one generation to the next, providing a potential mechanism for the inheritance of environmentally acquired traits. Transgenerational inheritance of RNA interference phenotypes in Caenorhabditis elegans provides an excellent model to study this phenomenon, and whilst studies have implicated both chromatin modifications and small RNA pathways in heritable silencing their relative contributions remain unclear. Here we demonstrate that the histone methyltransferases SET-25 and SET-32 are required for the establishment of a transgenerational silencing signal but not for long-term maintenance of this signal between subsequent generations, suggesting that transgenerational epigenetic inheritance is a multi-step process with distinct genetic requirements for establishment and maintenance of heritable silencing. Furthermore, small RNA sequencing reveals that the abundance of secondary siRNAs (thought to be the effector molecules of heritable silencing) does not correlate with silencing phenotypes. Together, our results suggest that the current mechanistic models of epigenetic inheritance are incomplete.

genetics↗