Biology subjects

Kramer, S.

Publications and source records attributed to Kramer, S..

7 recordsLinked to original sources

bioRxiv2021

OPEN leaf: an open-source cloud-based phenotyping system for tracking dynamic changes at leaf-specific resolution in Arabidopsis

The first draft of the Arabidopsis genome was released more than 20 years ago and despite intensive molecular research, more than 30% of Arabidopsis genes remained uncharacterized or without an assigned function. This is in part due to gene redundancy within gene families or the essential nature of genes, where their deletion results in lethality (i.e., the dark genome). High-throughput plant phenotyping (HTPP) offers an automated and unbiased approach to characterize subtle or transient phenotypes resulting from gene redundancy or inducible gene silencing; however, commercial HTPP platforms remain unaffordable. Here we describe the design and implementation of OPEN leaf, an open-source HTPP system with cloud connectivity and remote bilateral communication to facilitate data collection, sharing and processing. OPEN leaf, coupled with the SMART imaging processing package was able to consistently document and quantify dynamic morphological changes over time at the whole rosette level and also at leaf-specific resolution when plants experienced changes in nutrient availability. The modular design of OPEN leaf allows for additional sensor integration. Notably, our data demonstrate that VIS sensors remain underutilized and can be used in high-throughput screens to identify characterize previously unidentified phenotypes in a leaf-specific manner. Significance StatementMany bottlenecks exist in high-throughput phenotyping involving computing power for processing and a lack of focus on abiotic stresses that has prevented an advancement in phenotyping on par with genotyping. Therefore, we create an automated HTP system that performs nutrient studies on Arabidopsis thaliana with cloud-based image processing that quantifies plant traits at a whole and leaf-level.

plant biology↗

bioRxiv2021

The nucleolar DExD/H protein Hel66 is involved in ribosome biogenesis in Trypanosoma brucei

The biosynthesis of ribosomes is a complex cellular process involving ribosomal RNA, ribosomal proteins and several further trans-acting factors. DExD/H box proteins constitute the largest family of trans-acting protein factors involved in this process. Several members of this protein family have been directly implicated in ribosome biogenesis in yeast. In trypanosomes, ribosome biogenesis differs in several features from the process described in yeast. Here, we have identified the DExD/H box helicase Hel66 as being involved in ribosome biogenesis. The protein is unique to Kinetoplastida, localises to the nucleolus and its depletion via RNAi caused a severe growth defect. Loss of the protein resulted in a decrease of global translation and accumulation of rRNA processing intermediates for both the small and large ribosomal subunits. Only a few factors involved in trypanosome rRNA biogenesis have been described so far and our findings contribute to gaining a more comprehensive picture of this essential process.

microbiology↗

bioRxiv2021

Integrated cycles for urban biomass as a strategy to promote a CO2-neutral society - A feasibility study

Progressive global warming is one of the biggest challenges civilization is facing today. The establishment of a carbon dioxide (CO2)-neutral society based on sustainable value creation cycles is required to stop this development. The Integrated Cycles for Urban Biomass (ICU) concept is a new concept towards a CO2-neutral society. The integration of closed biomass cycles into residential buildings enable efficient resource utilization and avoid transport of biowaste. In this scenario, biowaste is degraded on-site into biogas that is converted into heat and electricity. The liquid fermentation residues are upgraded by nitrification processes (e.g., by a soiling(R)-process, EP3684909A1) to refined fertilizer, which can be used subsequently in house-internal gardens to produce fresh food for residents. Whereas this scenario sounds promising, comprehensive evaluations of produced amounts of biogas and food, saved CO2 and costs as well as social-cultural aspects are lacking. To assess these points, a feasibility study was performed, which estimated the material and energy flows based on simulations of the biogas process and food production. The calculations show that a residential complex with 100 persons can generate 21 % of the annual power (electrical and heat) consumption from the accumulated biowaste. The nitrogen (N) in the liquid fermentation residues enables the production of up to 6.3 t of fresh mass of lettuce per year in a 70 m2 professional hydroponic production area. The amount of produced lettuce corresponds to the amount of calories required to feed four persons for one year. Additionally, due to the reduction of biowaste transport and the in-house food and fertilizer production, 6 468 kg CO2-equivalent (CO2-eq) per year are saved compared to a conventional building. While the ICU concept is technically feasible, its costs are still 1.5 times higher than the revenues. However, the model predictions show that the ICU concept becomes economically feasible in case food prices further increase and ICU is implemented at larger scale, e.g.; at the district level. Finally, this study demonstrates that the ICU implementation can be a worthwile contribution towards a sustainable CO2-neutral society and enable to decrease the demand for agricultural land.

bioengineering↗

bioRxiv2021

A novel SNF2 ATPase complex in Trypanosoma brucei with a role in H2A.Z-mediated chromatin remodelling

A cascade of histone acetylation events with subsequent incorporation of a histone H2A variant plays an essential part in transcription regulation in various model organisms. A key player in this cascade is the chromatin remodelling complex SWR1, which replaces the canonical histone H2A with its variant H2A.Z. Transcriptional regulation of polycistronic transcription units in the unicellular parasite Trypanosoma brucei has been shown to be highly dependent on acetylation of H2A.Z, which is mediated by the histone-acetyltransferase HAT2. The chromatin remodelling complex which mediates H2A.Z incorporation is not known and an SWR1 orthologue in trypanosomes has not yet been reported. In this study, we identified and characterised an SWR1-like remodeller complex in T. brucei that is responsible for Pol II-dependent transcriptional regulation. Bioinformatic analysis of potential SNF2 DEAD/Box helicases, the key component of SWR1 complexes, identified a 1211 amino acids-long protein that exhibits key structural characteristics of the SWR1 subfamily. Systematic protein-protein interaction analysis revealed the existence of a novel complex exhibiting key features of an SWR1-like chromatin remodeller. RNAi-mediated depletion of the ATPase subunit of this complex resulted in a significant reduction of H2A.Z incorporation at transcription start sites and a subsequent decrease of steady-state mRNA levels. Furthermore, depletion of SWR1 and RNA-polymerase II (Pol II) caused massive chromatin condensation. The potential function of several proteins associated with the SWR1-like complex and with HAT2, the key factor of H2A.Z incorporation, is discussed. Author summaryTrypanosoma brucei is the causative agent of African trypanosomiasis (sleeping sickness) in humans and nagana in cattle. Its unusual genomic organisation featuring large polycistronic units requires a general mechanism of transcription initiation, because individual gene promoters are mostly absent. Despite the fact that the histone variant H2A.Z has previously been identified as a key player of transcription regulation, the complex responsible for correct H2A.Z incorporation at transcription start sites (TSS) remains elusive. In other eukaryotes, SWR1, a SNF2 ATPase-associated chromatin remodelling complex, is responsible for correct incorporation of this histone variant. This study identified a SWR1-like complex in T. brucei. Depletion of the SNF2 ATPase resulted in a reduction of H2A.Z incorporation at the TSS and decreased steady-state mRNA levels accompanied by chromatin condensation. In addition to the SWR1-like complex, we also identified a trypanosome-specific HAT2 complex that includes the histone acetyltransferases HAT2, a key player in the H2A.Z incorporation process. This complex has a trypanosome-specific composition that is different from the NuA4/TIP60 complex in Saccharomyces cerevisiae.

microbiology↗

bioRxiv2021

A metagenomic-based stufy of two sites from the Barbadian reef system

We study the microbiome of sea water collected from two locations of the Barbadian coral reefs. The two sites differ in several environmental and ecological variables including their endogenous benthic community in addition to their proximity to urban development and runoffs from inland watersheds. The composition of the microbial community was estimated using whole genome DNA shotgun sequencing. Although both sites exhibit a similar degree of richness, the less urbanized site (Maycocks reef at Hangmans Bay) is strongly concentrated with phototrophs. In comparison, the more urbanized location (Bellairs Research Institute) is enriched for copiotrophs, macroalgal symbionts and marine-related disease-bearing organisms from taxa scattered across the tree of life. Overall, our samples and associated measurements of chemical and environmental qualities of the water are in line with previous marine microbiome profiles of warm ocean surface waters. This suggests our approach captures salient information regarding the state of each coral reef site and sets the stage for larger longitudinal studies of coral reef dynamics in Barbados.

microbiology↗

bioRxiv2020

Is mRNA decapping activity of ApaH like phosphatases (ALPHs) the reason for the loss of cytoplasmic ALPHs in all eukaryotes but Kinetoplastida?

BackgroundApaH like phosphatases (ALPHs) originate from the bacterial ApaH protein and are present in eukaryotes of all eukaryotic super-groups; still, only two proteins have been functionally characterised. One is ALPH1 from the Kinetoplastid Trypanosoma brucei that we recently found to be the mRNA decapping enzyme of the parasite. mRNA decapping by ALPHs is unprecedented in eukaryotes, which usually use nudix hydrolases, but the bacterial ancestor protein ApaH was recently found to decap non-conventional caps of bacterial mRNAs. These findings prompted us to explore whether mRNA decapping by ALPHs is restricted to Kinetoplastida or more widespread among eukaryotes. ResultsWe screened 824 eukaryotic proteomes with a newly developed Python-based algorithm for the presence of ALPHs and used the data to refine phylogenetic distribution, conserved features, additional domains and predicted intracellular localisation of ALPHs. We found that most eukaryotes have either no ALPH (500/824) or very short ALPHs, consisting almost exclusively of the catalytic domain. These ALPHs had mostly predicted non-cytoplasmic localisations, often supported by the presence of transmembrane helices and signal peptides and in two cases (one in this study) by experimental data. The only exceptions were ALPH1 homologues from Kinetoplastida, that all have unique C-terminal and mostly unique N-terminal extension, and at least the T. brucei enzyme localises to the cytoplasm. Surprisingly, despite of these non-cytoplasmic localisations, ALPHs from all eukaryotic super-groups had in vitro mRNA decapping activity. ConclusionsALPH was present in the last common ancestor of eukaryotes, but most eukaryotes have either lost the enzyme since, or use it exclusively outside the cytoplasm in organelles in a version consisting of the catalytic domain only. While our data provide no evidence for the presence of further mRNA decapping enzymes among eukaryotic ALPHs, the broad substrate range of ALPHs that includes mRNA caps provides an explanation for the selection against the presence of a cytoplasmic ALPH protein as a mean to protect mRNAs from unregulated degradation. Kinetoplastida succeeded to exploit ALPH as their mRNA decapping enzyme, likely using the Kinetoplastida-unique N- and C-terminal extensions for regulation.

evolutionary biology↗

bioRxiv2020

Parallel monitoring of mRNA abundance, localisation and compactness with correlative single molecule FISH on LR White embedded samples

Single mRNA molecules are frequently detected by single molecule fluorescence in situ hybridisation (smFISH) using branched DNA technology. While providing strong and background-reduced signals, the method is inefficient in detecting mRNAs within dense structures, in monitoring mRNA compactness and in quantifying abundant mRNAs. To overcome these limitations, we have hybridised slices of high pressure frozen, LR White embedded cells (LR White smFISH). mRNA detection is physically restricted to the surface of the resin. This enables single molecule detection of RNAs with accuracy comparable to RNA sequencing, irrespective of their abundance, while at the same time providing spatial information on RNA localisation that can be complemented with immunofluorescence and electron microscopy, as well as electron tomography. Moreover, LR White embedding restricts the number of available probe pair recognition sites for each mRNA to a small subset. As a consequence, differences in signal intensities between RNA populations reflect differences in RNA tertiary structures, and we show that the method can be employed to probe for mRNA compactness. We apply LR White smFISH to answer some outstanding questions related to trans-splicing, RNA granules and mitochondrial RNA editing, using trypanosomes and their versatile RNA biology as a model system.

cell biology↗