Biology subjects

Haring, M.

Publications and source records attributed to Haring, M..

5 recordsLinked to original sources

bioRxiv2025

7β-hydroxysteroid dehydratase Hsh3 eliminates the 7-hydroxy group of the bile salt ursodeoxycholate during degradation by Sphingobium sp. strain Chol11 and other Sphingomonadaceae

Bile salts are steroids with distinctive hydroxylation patterns that are produced and excreted by vertebrates. In contrast to common human bile salts, ursodeoxycholate (UDCA) has a 7-hydroxy group in {beta}-configuration and is used in large amounts for treatment of diverse gastrointestinal diseases. We isolated the 7{beta}-hydroxysteroid dehydratase Hsh3 that is involved in UDCA degradation by Sphingobium sp. strain Chol11. Hsh3 eliminates the 7{beta}-hydroxy group as water, leading to a double bond in the B-ring. This is similar to 7-hydroxysteroid dehydratases in this and other strains, but Hsh3 is evolutionary different from these. Purified Hsh3 accepted steroids with and without side chain as substrates and had minor activity with 7-hydroxy groups. The deletion mutant strain Chol11 {Delta}hsh3 had impacted growth with UDCA and accumulated a novel compound. The compound was identified as 3,5-dihydroxy-H-methyl-hexahydro-5-indene-1-one-propanoate, consisting of steroid rings C and D with a C3-side chain carrying the former 7{beta}-hydroxy group, indicating that Hsh3 activity is important especially for the later stages of bile salt-degradation. Hsh3 homologs were found in other sphingomonads that were also able to degrade UDCA as well as in environmental metagenomes. Thus, Hsh3 adds to the bacterial enzyme repertoire for degrading a variety of differently hydroxylated bile-salts. ImportanceThe bacterial degradation of different bile salts is not only important for the removal of these steroidal compounds from the environment but also harbors interesting enzymes for steroid biotechnology. The 7{beta}-hydroxy bile salt UDCA naturally occurs in high concentration in the feces of black bears and has important pharmaceutical relevance for the treatment of different liver-related diseases, for which it is administered in high and increasing amounts. Additionally, it is present in the bile salt pool of humans in trace amounts. While ursodeoxycholate degradation is environmentally important, the enzyme Hsh3 modifies the hydroxy group that confers the medically relevant properties and thus might be interesting for microbiome analyses and biotechnological applications.

microbiology↗

bioRxiv2023

RdDM mutants reduce histone methylation rather than DNA methylation at the paramutated maize b1 enhancer

Paramutation is the transfer of mitotically and meiotically heritable silencing information between two alleles. With paramutation at the maize booster1 (b1) locus, the low expressed B epiallele heritably changes the high expressed B-I epiallele into B with 100% frequency. This requires specific tandem repeats and multiple components of the RNA-directed DNA methylation (RdDM) pathway, including the RNA-dependent RNA polymerase (RdRP, mediator of paramutation1, MOP1), the second-largest subunit of RNA polymerase IV and V (NRP(D/E)2a, mediator of paramutation2, MOP2), and the largest subunit of RNA Polymerase IV (NRPD1, mediator of paramutation3, MOP3). Mutations in mop genes prevent paramutation and release silencing at the B epiallele. In this study we investigated the effect of mutations in mop1, mop2 and mop3 on chromatin structure and DNA methylation at the B epiallele, and especially the regulatory hepta-repeat 100 kb upstream of the b1 gene. We show that mutations in mop1 and mop3 result in decreased repressive histone modifications H3K9me2 and H3K27me2 at the hepta-repeat. Associated with this decrease are partial activation of the hepta-repeat enhancer function, formation of a multi-loop structure, and elevated b1 expression. In mop2 mutants, which do not show elevated b1 expression, H3K9me2 and H3K27me2 and a single-loop structure like in wild type B are retained. Surprisingly, high DNA methylation levels at the B hepta-repeat remains in all three mutants. Our results raise the possibility of MOP factors mediating RNA-directed histone methylation rather than RNA-directed DNA methylation at the b1 locus.

plant biology↗

bioRxiv2023

Ectopic Expression of Distinct PLC Genes Identifies 'Compactness' as Novel Architectural Shoot Strategy to Cope with Drought Stress

Phospholipase C (PLC) has been implicated in several stress responses, including drought. Overexpression (OE) of PLC has been shown to improve drought tolerance in various plant species. Arabidopsis contains nine PLC genes, subdivided into four clades. Earlier, OE of PLC3, -5 or -7 were found to increase Arabidopsis drought tolerance. Here, we confirm this for three other PLCs: PLC2, the only constitutively expressed AtPLC; PLC4, reported to have reduced salt tolerance; and PLC9, of which the encoded enzyme was presumed to be catalytically inactive. To compare each PLC and to discover any other potential phenotype, two independent OE lines of six AtPLC genes, representing all four clades, were simultaneously monitored with the GROWSCREEN FLUORO phenotyping platform, under both control- and mild drought conditions. To investigate which tissues were most relevant to achieve drought survival, we additionally expressed AtPLC5 using 13 different cell- or tissue-specific promoters. While no significant differences in plant size, biomass or photosynthesis were found between PLC lines and wild-type (WT) plants, all PLC-OE lines, as well as those tissue-specific lines that promoted drought survival, exhibited a stronger decrease in convex hull perimeter (= increase in compactness) under water deprivation compared to WT. Increased compactness has not been associated with drought or decreased water loss before, though a hyponastic decrease in compactness in response to increased temperatures has been associated with water loss. We pose that increased compactness leads to decreased water loss and potentially provides a new breeding trait to select for drought tolerance.

plant biology↗

bioRxiv2023

Polychaetoid/ZO-1 strengthens cell junctions under tension while localizing differently than core adherens junction proteins

During embryonic development dramatic cell shape changes and movements re-shape the embryonic body plan. These require robust but dynamic linkage between the cell-cell adherens junctions and the force-generating actomyosin cytoskeleton. Our view of this linkage has evolved, and we now realize linkage is mediated by a mechanosensitive multiprotein complex assembled via multivalent connections. Here we combine genetic, cell biological and modeling approaches to define the mechanism of action and functions of an important player, Drosophila Polychaetoid, homolog of mammalian ZO-1. Our data reveal that Pyd reinforces cell junctions under elevated tension, and facilitates cell rearrangements. Pyd is important to maintain junctional contractility and in its absence cell rearrangements stall. We next use structured illumination microscopy to define the molecular architecture of cell-cell junctions during these events. The cadherin-catenin complex and Cno both localize to puncta along the junctional membrane, but are differentially enriched in different puncta. Pyd, in contrast, exhibits a distinct localization to strands that extend out from the region occupied by core junction proteins. We then discuss the implications for the protein network at the junction-cytoskeletal interface, suggesting different proteins localize and function in distinct ways but combine to produce robust connections.

cell biology↗

bioRxiv2018

Molecular architecture of the mouse nervous system

The mammalian nervous system executes complex behaviors controlled by specialised, precisely positioned and interacting cell types. Here, we used RNA sequencing of half a million single cells to create a detailed census of cell types in the mouse nervous system. We mapped cell types spatially and derived a hierarchical, data-driven taxonomy. Neurons were the most diverse, and were grouped by developmental anatomical units, and by the expression of neurotransmitters and neuropeptides. Neuronal diversity was driven by genes encoding cell identity, synaptic connectivity, neurotransmission and membrane conductance. We discovered several distinct, regionally restricted, astrocytes types, which obeyed developmental boundaries and correlated with the spatial distribution of key glutamate and glycine neurotransmitters. In contrast, oligodendrocytes showed a loss of regional identity, followed by a secondary diversification. The resource presented here lays a solid foundation for understanding the molecular architecture of the mammalian nervous system, and enables genetic manipulation of specific cell types.

neuroscience↗