Biology subjects

Hiller, M.

Publications and source records attributed to Hiller, M..

At least 37 records · Page 2Linked to original sources

bioRxiv2023

RETINOID ORPHAN RECEPTOR GAMMA T (RORgT) PROMOTES INFLAMMATORY EOSINOPHILIA BUT IS DISPENSABLE FOR INNATE IMMUNE-MEDIATED COLITIS

Inflammatory bowel diseases (IBD) result from uncontrolled inflammation in the intestinal mucosa leading to damage and loss of function. Both innate and adaptive immunity contribute to the inflammation of IBD and innate and adaptive immune cells reciprocally activate each other in a forward feedback loop. In order to better understand innate immune contributions to IBD, we developed a model of spontaneous 100% penetrant, early onset colitis that occurs in the absence of adaptive immunity by crossing villin-TNFAIP3 mice to RAG1-/- mice (TRAG mice). This model is driven by microbes and features increased levels of innate lymphoid cells in the intestinal mucosa. To investigate the role of type 3 innate lymphoid cells (ILC3) in the innate colitis of TRAG mice, we crossed them to retinoid orphan receptor gamma t deficient (Ror{gamma}t-/-) mice. Ror{gamma}t-/- x TRAG mice exhibited markedly reduced eosinophilia in the colonic mucosa, but colitis persisted in these mice. Colitis in Ror{gamma}t-/- x TRAG mice was characterized by increased infiltration of the intestinal mucosa by neutrophils, inflammatory monocytes, macrophages and other innate cells. RNA and cellular profiles of Ror{gamma}t-/- x TRAG mice were consistent with a lack of ILC3 and ILC3 derived cytokines, reduced antimicrobial factors, increased activation oof epithelial repair processes and reduced activation of epithelial cell STAT3. The colitis in Ror{gamma}t-/- x TRAG mice was ameliorated by antibiotic treatment indicating that microbes contribute to the ILC3-independent colitis of these mice. Thus, Ror{gamma}t promotes eosinophilia but Ror{gamma}t and Ror{gamma}t-dependent ILC3 are dispensable for the innate colitis in TRAG mice.

immunology↗

bioRxiv2023

Structure-function relationships underpin disulfide loop cleavage-dependent activation of Legionella pneumophila lysophosholipase A PlaA

Legionella pneumophila, the causative agent of a life-threatening pneumonia, intracellularly replicates in a specialized compartment in lung macrophages, the Legionella-containing vacuole (LCV). Secreted proteins of the pathogen govern important steps in the intracellular life cycle including bacterial egress. Among these is the type II secreted PlaA which, together with PlaC and PlaD, belongs to the GDSL phospholipase family found in L. pneumophila. PlaA shows lysophospholipase A (LPLA) activity which increases after secretion and subsequent processing by the zinc metalloproteinase ProA at residue E266/L267 located within a disulfide loop. Activity of PlaA contributes to the destabilization of the LCV in the absence of the type IVB-secreted effector SdhA. We here present the 3D structure of PlaA which shows a typical /{beta} hydrolase fold and reveals that the uncleaved disulfide loop forms a lid structure covering the catalytic triad S30/D278/H282. This leads to reduction of both substrate access and membrane interaction before activation; however, the catalytic and membrane interaction site gets more accessible when the disulfide loop is processed. After structural modelling, a similar activation process is suggested for the GDSL hydrolase PlaC, but not for PlaD. Furthermore, the size of the PlaA substrate binding site indicated preference towards phospholipids comprising ~16 carbon fatty acid residues which was verified by lipid hydrolysis, suggesting a molecular ruler mechanism. Indeed, mutational analysis changed the substrate profile with respect to fatty acid chain length. In conclusion, our analysis revealed the structural basis for the regulated activation and substrate preference of PlaA.

biochemistry↗

bioRxiv2023

Chiropterans are a hotspot for horizontal transfer of DNA transposons in Mammalia

Horizontal transfer of transposable elements is an important mechanism contributing to genetic diversity and innovation. Bats (order Chiroptera) have repeatedly been shown to experience horizontal transfer of transposable elements at what appears to be a high rate compared to other mammals. We investigated the occurrence of horizontally transferred DNA transposons involving bats. We found over 200 putative horizontally transferred elements within bats; sixteen transposons were shared across distantly related mammalian clades and two other elements were shared with a fish and two lizard species. Our results indicate that bats are a hotspot for horizontal transfer of DNA transposons. These events broadly coincide with the diversification of several bat clades, supporting the hypothesis that DNA transposon invasions have contributed to genetic diversification of bats.

molecular biology↗

bioRxiv2022

Bat pluripotent stem cells reveal unique entanglement between host and viruses

Bats have evolved features unique amongst mammals, including flight, laryngeal echolocation, and certain species have been shown to have a unique immune response that may enable them to tolerate viruses such as SARS-CoVs, MERS-CoVs, Nipah, and Marburg viruses. Robust cellular models have yet to be developed for bats, hindering our ability to further understand their special biology and handling of viral pathogens. To establish bats as new model study species, we generated induced pluripotent stem cells (iPSCs) from a wild greater horseshoe bat (Rhinolophus ferrumequinum) using a modified Yamanaka protocol. Rhinolophids are amongst the longest living bat species and are asymptomatic carriers of coronaviruses, including one of the viruses most closely related to SARS-CoV-2. Bat induced pluripotent stem (BiPS) cells were stable in culture, readily differentiated into all three germ layers, and formed complex embryoid bodies, including organoids. The BiPS cells were found to have a core pluripotency gene expression program similar to that of other species, but it also resembled that of cells attacked by viruses. The BiPS cells produced a rich set of diverse endogenized viral sequences and in particular retroviruses. We further validated our protocol by developing iPS cells from an evolutionary distant bat species Myotis myotis (greater mouse-eared bat) non-lethally sampled in the wild, which exhibited similar attributes to the greater horseshoe bat iPS cells, suggesting that this unique pluripotent state evolved in the ancestral bat lineage. Although previous studies have suggested that bats have developed powerful strategies to tame their inflammatory response, our results argue that they have also evolved mechanisms to accommodate a substantial load of endogenous viral sequences and suggest that the natural history of bats and viruses is more profoundly intertwined than previously thought. Further study of bat iPS cells and their differentiated progeny should advance our understanding of the role bats play as virus hosts, provide a novel method of disease surveillance, and enable the functional studies required to ascertain the molecular basis of bats unique traits.

developmental biology↗

bioRxiv2022

Integrating gene annotation with orthology inference at scale

Annotating coding genes and inferring orthologs are two classical challenges in genomics and evolutionary biology that have traditionally been approached separately, limiting scalability. We present TOGA, a method that integrates structural gene annotation and orthology inference. TOGA implements a different paradigm to infer orthologous loci, improves ortholog detection and annotation of conserved genes compared to state-of-the-art methods, and handles even highly-fragmented assemblies. TOGA scales to hundreds of genomes, which we demonstrate by applying it to 488 placental mammal and 501 bird assemblies, creating the largest comparative gene resources so far. Additionally, TOGA detects gene losses, enables selection screens, and automatically provides a superior measure of mammalian genome quality. Together, TOGA is a powerful and scalable method to annotate and compare genes in the genomic era.

genomics↗

bioRxiv2022

High quality genomes corroborate 29 chromosomes of the haploid Hyles (Lepidoptera: Sphingidae) karyotype

BackgroundMorphological and traditional genetic studies of the young Pliocene genus Hyles have led to the understanding that despite its importance for taxonomy, phenotypic similarity of wing patterns does not correlate with phylogenetic relationship. To gain insights into various aspects of speciation in the Spurge Hawkmoth (Hyles euphorbiae), we assembled a chromosome-level genome and investigated some of its characteristics. ResultsThe genome of a male H. euphorbiae was sequenced using PacBio and Hi-C data, yielding a 504 Mb assembly (scaffold N50 of 18.2 Mb) with 99.9% of data represented by the 29 largest scaffolds forming the haploid chromosome set. Consistent with this, FISH analysis of the karyotype revealed n = 29 chromosomes and a WZ/ZZ (female/male) sex chromosome system. Estimates of chromosome length based on the karyotype image provided an additional quality metric of assembled chromosome size. Rescaffolding the published male H. vespertilio genome resulted in a high-quality assembly (651 Mb, scaffold N50 of 22 Mb) with 98% of sequence data in the 29 chromosomes. The larger genome size of H. vespertilio (average 1C DNA value of 562 Mb) was accompanied by a proportional increase in repeats from 45% in H. euphorbiae (measured as 472 Mb) to almost 55% in H. vespertilio. Several wing pattern genes were found on the same chromosomes in the two species, with varying amounts and positions of repetitive elements and inversions possibly corrupting their function. ConclusionsOur two-fold comparative genomics approach revealed high gene synteny of the Hyles genomes to other Sphingidae and high correspondence to intact Merian elements, the ancestral linkage groups of Lepidoptera, with the exception of three simple fusion events. We propose a standardized approach for genome taxonomy using nucleotide homology via scaffold chaining as the primary tool combined with Oxford plots based on Merian elements to infer and visualize directionality of chromosomal rearrangements. The identification of wing pattern genes promises future understanding of the evolution of forewing patterns in the genus Hyles, although further sequencing data from more individuals are needed. The genomic data obtained provide additional reliable references for further comparative studies in hawkmoths (Sphingidae).

genomics↗

bioRxiv2022

Vision-related convergent gene losses reveal SERPINE3's unknown role in the eye

Despite decades of research, knowledge about the genes that are important for development and function of the mammalian eye and are involved in human eye disorders remains incomplete. During mammalian evolution, mammals that naturally exhibit poor vision or regressive eye phenotypes have independently lost many eye-related genes. This provides an opportunity to predict novel eye-related genes based on specific evolutionary gene loss signatures. Building on these observations, we performed a genome-wide screen across 49 mammals for functionally uncharacterized genes that are preferentially lost in species exhibiting lower visual acuity values. The screen uncovered several genes, including SERPINE3, a putative serine proteinase inhibitor. A detailed investigation of 381 additional mammals revealed that SERPINE3 is independently lost in 18 lineages that typically do not primarily rely on vision, predicting a vision-related function for this gene. To test this, we show that SERPINE3 has the highest expression in eyes of zebrafish and mouse. In the zebrafish retina, serpine3 is expressed in Mueller glia cells, a cell type essential for survival and maintenance of the retina. A CRISPR-mediated knockout of serpine3 in zebrafish resulted in alterations in eye shape and defects in retinal layering. Furthermore, two human polymorphisms that are in linkage with SERPINE3 are associated with eye-related traits. Together, these results suggest that SERPINE3 has a role in vertebrate eyes. More generally, by integrating comparative genomics with experiments in model organisms, we show that screens for specific phenotype-associated gene signatures can predict functions of uncharacterized genes.

evolutionary biology↗

bioRxiv2022

Nematodes can survive in a suspended form of life for indefinite time

Some organisms in nature have developed the ability to enter a state of suspended metabolism called cryptobiosis1 when environmental conditions are unfavorable. This state-transition requires execution of a combination of genetic and biochemical pathways1,2,3 that enable the organism to survive for prolonged periods. Recently, nematode individuals have been reanimated from Siberian permafrost after remaining in cryptobiosis. Preliminary analysis indicates that these nematodes belong to the genera Panagrolaimus and Plectus4. Here, we present precise radiocarbon dating indicating that the Panagrolaimus individuals have remained in cryptobiosis since the late Pleistocene ([~]46,000 years). Phylogenetic inference based on our genome assembly and a detailed morphological analysis demonstrate that they belong to an undescribed species, which we named Panagrolaimus n. sp. Comparative genome analysis revealed that the molecular toolkit for cryptobiosis in Panagrolaimus n. sp. and in C. elegans is partly orthologous. We show that biochemical mechanisms employed by these two species to survive desiccation and freezing under laboratory conditions are similar. Our experimental evidence also reveals that C. elegans dauer larvae can remain viable for longer periods in suspended animation than previously reported. Altogether, our findings demonstrate that nematodes evolved mechanisms potentially allowing them to suspend life over geological time scales.

genomics↗

bioRxiv2021

A haplotype-resolved genome assembly of the Nile rat facilitates exploration of the genetic basis of diabetes

The Nile rat (Avicanthis niloticus) is an important animal model for biomedical research, including the study of diurnal rhythms and type 2 diabetes. Here, we report a 2.5 Gb, chromosome-level reference genome assembly with fully resolved parental haplotypes, generated with the Vertebrate Genomes Project (VGP). The assembly is highly contiguous, with contig N50 of 11.1 Mb, scaffold N50 of 83 Mb, and 95.2% of the sequence assigned to chromosomes. We used a novel workflow to identify 3,613 segmental duplications and quantify duplicated genes. Comparative analyses revealed unique genomic features of the Nile rat, including those that affect genes associated with type 2 diabetes and metabolic dysfunctions. These include 14 genes that are heterozygous in the Nile rat or highly diverged from the house mouse. Our findings reflect the exceptional level of genomic detail present in this assembly, which will greatly expand the potential of the Nile rat as a model organism for genetic studies.

genomics↗

bioRxiv2021

Gene losses in the common vampire bat illuminate molecular adaptations to blood feeding

Feeding exclusively on blood, vampire bats represent the only obligate sanguivorous lineage among mammals. To uncover genomic changes associated with adaptations to this unique dietary specialization, we generated a new haplotype-resolved reference-quality genome of the common vampire bat (Desmodus rotundus) and screened 26 bat species for genes that were specifically lost in the vampire bat lineage. We discovered previously-unknown gene losses that relate to metabolic and physiological changes, such as reduced insulin secretion (FFAR1, SLC30A8), limited glycogen stores (PPP1R3E), and a distinct gastric physiology (CTSE). Other gene losses likely reflect the biased nutrient composition (ERN2, CTRL) and distinct pathogen diversity of blood (RNASE7). Interestingly, the loss of REP15 likely helped vampire bats to adapt to high dietary iron levels by enhancing iron excretion and the loss of the 24S-hydroxycholesterol metabolizing enzyme CYP39A1 could contribute to their exceptional cognitive abilities. Finally, losses of key cone phototransduction genes (PDE6H, PDE6C) suggest that these strictly-nocturnal bats completely lack cone-based vision. These findings enhance our understanding of vampire bat biology and the genomic underpinnings of adaptations to sanguivory.

evolutionary biology↗

bioRxiv2021

DENTIST - using long reads to close assembly gaps at high accuracy

BackgroundLong sequencing reads allow increasing contiguity and completeness of fragmented, short-read based genome assemblies by closing assembly gaps, ideally at high accuracy. While several gap closing methods have been developed, these methods often close an assembly gap with sequence that does not accurately represent the true sequence. FindingsHere, we present DENTIST, a sensitive, highly accurate and automated pipeline method to close gaps in short-read assemblies with long error-prone reads. DENTIST comprehensively determines repetitive assembly regions to identify reliable and unambiguous alignments of long reads to the correct loci, integrates a consensus sequence computation step to obtain a high base accuracy for the inserted sequence, and validates the accuracy of closed gaps. Unlike previous benchmarks, we generated test assemblies that have gaps at the exact positions where real short-read assemblies have gaps. Generating such realistic benchmarks for Drosophila (134 Mb genome), Arabidopsis (119 Mb), hummingbird (1 Gb) and human (3 Gb) and using simulated or real PacBio continuous long reads, we show that DENTIST consistently achieves a substantially higher accuracy compared to previous methods, while having a similar sensitivity. ConclusionDENTIST provides an accurate approach to improve the contiguity and completeness of fragmented assemblies with long reads. DENTISTs source code including a Snakemake workflow, conda package and Docker container is available at https://github.com/a-ludi/dentist. All test assemblies as a resource for future benchmarking are at https://bds.mpi-cbg.de/hillerlab/DENTIST/.

bioinformatics↗

bioRxiv2021

Phenotyping in the era of genomics: MaTrics a digital character matrix to document mammalian phenotypic traits coded numerically

A new and uniquely structured matrix of mammalian phenotypes, MaTrics (Mammalian Traits for Comparative Genomics) is presented in a digital form. By focussing on mammalian species for which genome assemblies are available, MaTrics provides an interface between mammalogy and comparative genomics. MaTrics was developed as part of a project to link phenotypic differences between mammals to differences in their genomes using Forward Genomics. Apart from genomes this approach requires information on homologous phenotypes that are numerically encoded (presence-absence; multistate character coding*) in a matrix. MaTrics provides these data, links them to at least one reference (e.g., literature, photographs, histological sections, CT-scans, or museum specimens) and makes them available in a machine actionable NEXUS-format. By making the data computer readable, MatTrics opens a new way for digitizing collections. Currently, MaTrics covers 147 mammalian species and includes 207 characters referring to structure, morphology, physiology, ecology and ethology. Researching these traits revealed substantial knowledge gaps, highlighting the need for substantial phenotyping efforts in the genomic era. Using the trait information documented in MaTrics, previous Forward Genomics screens identified changes in genes that are associated with various phenotypes, ranging from fully-aquatic lifestyle to dietary specializations. These results motivate the continuous expansion of phenotype information, both by filling research gaps or by adding additional taxa and traits. MaTrics is digitally available online within the data repository Morph{middle dot}D{middle dot}Base (www.morphdbase.de).

zoology↗

bioRxiv2021

Determining pathogenicity of variants of uncertain significance and identification of a founder variant in the epilepsy-associated gene, SZT2

Biallelic pathogenic variants in SZT2 result in a neurodevelopmental disorder with shared features, including early-onset epilepsy, developmental delay, macrocephaly, and corpus callosum abnormalities. SZT2 is as a critical scaffolding protein in the amino acid sensing arm of the mTOR signaling pathway. Due to its large size (3432 amino acids), lack of crystal structure, and absence of functional domains, it is difficult to determine the pathogenicity of SZT2 missense and in-frame deletions. We report a cohort of twelve individuals with biallelic SZT2 variants and phenotypes consistent with SZT2-related neurodevelopmental disorder. The majority of this cohort contained one or more SZT2 variants of uncertain significance (VUS). We developed a novel individualized platform to functionally characterize SZT2 VUSs. We identified a recurrent in-frame deletion (SZT2 p.Val1984del) which was determined to be a loss-of-function variant and therefore likely pathogenic. Haplotype analysis determined this single in-frame deletion is a founder variant in those of Ashkenazi Jewish ancestry. Overall, we present a FACS-based rapid assay to distinguish pathogenic variants from VUSs in SZT2, using an approach that is widely applicable to other mTORopathies including the most common causes of the focal genetic epilepsies, DEPDC5, TSC1/2, MTOR and NPRL2/3.

genetics↗

bioRxiv2020

Genomic and anatomical comparisons of skin support independent adaptation to life in water by cetaceans and hippos

The macroevolutionary transition from terra firma to obligatory inhabitance of the marine hydrosphere has occurred twice in the history of Mammalia: Cetacea and Sirenia. In the case of Cetacea (whales, dolphins, porpoises), molecular phylogenies provide unambiguous evidence that fully aquatic cetaceans and semiaquatic hippopotamids (hippos) are each others closest living relatives. Ancestral reconstructions further suggest that some adaptations to the aquatic realm evolved in the common ancestor of Cetancodonta (Cetacea+Hippopotamidae). An alternative hypothesis is that these adaptations evolved independently in cetaceans and hippos. Here, we focus on the integumentary system and evaluate these hypotheses by integrating new histological data for cetaceans and hippos, the first genome-scale data for pygmy hippopotamus, and comprehensive genomic screens and molecular evolutionary analyses for protein-coding genes that have been inactivated in hippos and cetaceans. We identified ten skin-related genes that are inactivated in both cetaceans and hippos, including genes that are related to sebaceous glands, hair follicles, and epidermal differentiation. However, none of these genes exhibit inactivating mutations that are shared by cetaceans and hippos. Mean dates for the inactivation of skin genes in these two clades serve as proxies for phenotypic changes and suggest that hair reduction/loss, the loss of sebaceous glands, and changes to the keratinization program occurred ~16 million years earlier in cetaceans (~46.5 Ma) than in hippos (~30.5 Ma). These results, together with histological differences in the integument and prior analyses of oxygen isotopes from stem hippopotamids ("anthracotheres"), support the hypothesis that aquatic adaptations evolved independently in hippos and cetaceans.

evolutionary biology↗

bioRxiv2020

Filaggrin deficient mice have a lower threshold for cutaneous allergen sensitization but do not develop spontaneous skin inflammation or atopy

Defects of filaggrin (FLG) compromise epidermal barrier function and represent an important known genetic risk factor for atopic dermatitis (AD), but also for systemic atopy, including allergic sensitization and asthma. The flaky tail mouse model, widely used to address mechanisms of atopy induction by barrier-defective skin, harbors two mutations that affect the skin barrier, the mutation Flgft, resulting in near-complete loss of FLG expression, and the matted mutation inactivating transmembrane protein 79 (Tmem79). Upon separation of the two mutant loci, which are closely linked on chromosome 3, mice defective only for Tmem79 featured pronounced dermatitis and systemic atopy. Upon extensive backcross to BALB/c, also Flgft/ft mice (assumed to be wild type for Tmem79), developed AD-like dermatitis and reproduced the human atopic march, with high IgE levels and spontaneous asthma, suggesting a key role for functional Flg in protection from atopy also in mice. In contrast, BALB/c mice congenic for a targeted Flg knock out mutation did not develop skin inflammation or atopy. To resolve this discrepancy, we generated Flg-deficient mice on a pure BALB/c background by inactivating the Flg gene in BALB/c embryos. These animals feature an ichthyosis phenotype, but do not develop spontaneous dermatitis or systemic atopy. We sequenced the genome of the atopic Flgft BALB/c congenics and discovered that they were unexpectedly homozygous for the atopy-causing Tmem79matted mutation. In summary, we show that Flg-deficiency does not cause atopy in mice. This finding is in line with lack of atopic disease in a fraction of Ichthyosis vulgaris patients carrying two FLG null alleles. However, absence of FLG may promote and modulate dermatitis caused by other genetic barrier defects, as skin inflammation in Tmem79ma/maFlgft/ft BALB/c congenics is qualitatively different compared to Tmem79ma/ma mice.

immunology↗

bioRxiv2020

Algorithmic Assessment of Missense Mutation Severity in the Von-Hippel Lindau Protein

Von Hippel-Lindau disease (VHL) is an autosomal dominant rare disease that causes the formation of angiogenic tumors. When functional, pVHL acts as an E3 ubiquitin ligase that negatively regulates hypoxia inducible factor (HIF). Genetic mutations that perturb the structure of pVHL result in dysregulation of HIF, causing a wide array of tumor pathologies including retinal angioma, pheochromocytoma, central nervous system hemangioblastoma, and clear cell renal carcinoma. These VHL-related cancers occur throughout the lifetime of the patient, requiring frequent intervention procedures, such as surgery, to remove the tumors. Although VHL is classified as a rare disease (1 in 39,000 to 1 in 91,000 affected) there is a large heterogeneity in genetic mutations listed for observed pathologies. Understanding how these specific mutations correlate with the myriad of observed pathologies for VHL could provide clinicians insight into the potential severity and onset of disease. Using a set of 285 ClinVar mutations in VHL, we developed a multiparametric scoring algorithm to evaluate the overall clinical severity of missense mutations in pVHL. The mutations were assessed according to eight weighted parameters as a comprehensive evaluation of protein misfolding and malfunction. Higher mutation scores were strongly associated with pathogenicity. Our approach establishes a novel in silico method by which VHL-specific mutations can be assessed for their severity and effect on the biophysical functions of the VHL protein.View Full Text

pathology↗

bioRxiv2020

Broad Host Range of SARS-CoV-2 Predicted by Comparative and Structural Analysis of ACE2 in Vertebrates

The novel coronavirus SARS-CoV-2 is the cause of Coronavirus Disease-2019 (COVID-19). The main receptor of SARS-CoV-2, angiotensin I converting enzyme 2 (ACE2), is now undergoing extensive scrutiny to understand the routes of transmission and sensitivity in different species. Here, we utilized a unique dataset of 410 vertebrates, including 252 mammals, to study cross-species conservation of ACE2 and its likelihood to function as a SARS-CoV-2 receptor. We designed a five-category ranking score based on the conservation properties of 25 amino acids important for the binding between receptor and virus, classifying all species from very high to very low. Only mammals fell into the medium to very high categories, and only catarrhine primates in the very high category, suggesting that they are at high risk for SARS-CoV-2 infection. We employed a protein structural analysis to qualitatively assess whether amino acid changes at variable residues would be likely to disrupt ACE2/SARS-CoV-2 binding, and found the number of predicted unfavorable changes significantly correlated with the binding score. Extending this analysis to human population data, we found only rare (<0.1%) variants in 10/25 binding sites. In addition, we observed evidence of positive selection in ACE2 in multiple species, including bats. Utilized appropriately, our results may lead to the identification of intermediate host species for SARS-CoV-2, justify the selection of animal models of COVID-19, and assist the conservation of animals both in native habitats and in human care.

genomics↗

bioRxiv2020

Recapitulating evolutionary divergence in a single regulatory element causes expression changes of the lens gene Tdrd7

Mutations in cis-regulatory elements play important roles for phenotypic changes during evolution. Eye degeneration in the blind mole rat (BMR) and other subterranean mammals is significantly associated with widespread divergence of eye regulatory elements, but the effect of these regulatory mutations on eye development and function has not been explored. Here, we investigate the effect of mutations observed in the BMR sequence of a conserved non-coding element upstream of Tdrd7, a pleiotropic gene required for lens development and spermatogenesis. We first show that this conserved element is a transcriptional repressor in lens cells and that the BMR sequence partially lost repressor activity. Next, we recapitulated the evolutionary changes by precisely replacing the endogenous regulatory element in a mouse line by the orthologous BMR sequence with CRISPR-Cas9. Strikingly, this repressor element has a large effect, causing a more than two-fold up-regulation of Tdrd7 in developing lens. Interestingly, the increased mRNA level does not result in a corresponding increase in TDRD7 protein nor an obvious lens phenotype, likely explained by buffering at the posttranscriptional level. Our results are consistent with eye degeneration in subterranean mammals having a polygenic basis where many small-effect mutations in different eye-regulatory elements collectively contribute to phenotypic differences.

evolutionary biology↗