Biology subjects

Menke, D. B.

Publications and source records attributed to Menke, D. B..

9 recordsLinked to original sources

bioRxiv2025

Developmental and transcriptional programs that define the initiation of the forelimb

The vertebrate forelimb initiates as a localized swelling in the somatic lateral plate mesoderm (somatic LPM) in response to TBX5-dependent transcription. The molecular pathways driving limb morphogenesis have been extensively studied but the steps directly preceding limb bud formation remain poorly characterized. To address this, we defined the temporal onset of forelimb initiation in mouse embryos using sequencing based high-throughput approaches (RNA-seq, scRNA-seq, ChIP-seq, and Ribo-ITP) benchmarked to known features in forelimb development, identifying four distinct stages. Using scRNA-seq at the onset of forelimb-specific transcription, we determined the transcriptional profile of the somatic LPM and identified signature genes that distinguish the nascent forelimb from other cell types. This group includes multiple genes involved in neural projection as well as cell adhesion. Interestingly, these genes are highly enriched for TBX5 binding sites, suggesting they are candidate early transcriptional targets of TBX5. As TBX5 is essential for forelimb outgrowth, the identification of these genes suggests new mechanistic models for TBX5-driven limb initiation.

developmental biology↗

bioRxiv2023

A new cell culture resource for investigations of reptilian gene function

The recent establishment of CRISPR/Cas9 gene editing in A. sagrei lizards makes it a powerful model system for studies of reptilian gene function. To enhance the versatility of this model, we developed an immortalized lizard fibroblast cell line (ASEC-1) for the exploration of reptilian gene function in cellular processes. We demonstrate the use of this in vitro system by scrutinizing the role of primary cilia in lizard Hedgehog (Hh) signaling. Through CRISPR/Cas9 mutagenesis we disrupted the ift88 gene, which is required for ciliogenesis in diverse organisms. We find that the loss of itf88 from lizard cells results in an absence of primary cilia, a partial derepression of gli1 transcription, and an inability of the cells to respond to the Smoothened agonist, SAG. Through a cross-species analysis of SAG-induced transcriptional responses in cultured limb bud cells, we further determined that [~]46% of genes induced as a response to Hh pathway activation in A. sagrei, are also SAG-responsive in M. musculus limb bud cells. Our results highlight conserved and diverged aspects of Hh signaling in anoles and establish a new resource for investigations of reptilian gene function.

developmental biology↗

bioRxiv2023

Maturation and Refinement of the Maculae and Foveae in the Anolis sagrei lizard

In primates, the fovea is a pit in the center of the macula, which is a region of the retina with a high concentration of cone photoreceptor cells that accounts for a large degree of visual acuity in primates. The maturation of this primate visual acuity area is characterized by the shallowing and widening of the foveal pit, a decrease in the diameter of the rod-free zone, and packing of photoreceptor cells more densely into this area of the retina, which occurs sometime after birth. Maturation occurs concurrently with progressing age and development, which further correlates with increasing eye size, retinal length, and retinal area. These observations have led to the hypothesis that the maturation of the fovea might be a function of mechanical variables that remodel the retina. However, this has never been explored outside of primates. Here, we take advantage of the Anolis sagrei lizard, which has a bifoveated retina, to study maturation of the fovea and macula. Eyes were collected from male and female lizards from five different age groups - hatchling, 2-month, 4-month, 6-month, and adult. We found that Anolis maculae undergo a maturation process somewhat different than what has been observed in primates. Anole macular diameters actually increase in size and undergo minimal photoreceptor cell packing, possessing a near complete complement of these cells at the time of hatching. As the anole eye expands, foveal centers experience little change in overall retina cell density with most cell redistribution occurring at the macular borders and peripheral retina areas. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/529081v1_ufig1.gif" ALT="Figure 1"> View larger version (73K): org.highwire.dtl.DTLVardef@effd8dorg.highwire.dtl.DTLVardef@1ef65b3org.highwire.dtl.DTLVardef@1fde14dorg.highwire.dtl.DTLVardef@d2c0b2_HPS_FORMAT_FIGEXP M_FIG C_FIG Highlights- Foveal and retinal refinement occurs post-hatching into adulthood in anoles. - The temporal pit shallows and widens, whereas central pit morphology remains relatively constant with age. - Retina cell number within a section decreases as the eye grows larger with age. - Anole macular diameter increases from hatching to adult but does not increase proportionately to overall eye growth.

developmental biology↗

bioRxiv2021

Development and retinal remodeling in the brown anole lizard (Anolis sagrei)

BackgroundThe fovea, a pit in the retina, is believed to be important for high-acuity vision and is a feature found in the eyes of humans and a limited number of vertebrate species that include certain primates, birds, lizards, and fish. At present, model systems currently used for ocular research lack a foveated retina and studies investigating fovea development have largely been limited to histological and molecular studies in primates. As a result, progress towards understanding the mechanisms involved in regulating fovea development in humans is limited and is completely lacking in other, non-primate, vertebrates. To address this knowledge gap, we provide here a detailed histological atlas of retina and fovea development in the bifoveated Anolis sagrei lizard, a novel reptile model for fovea research. We also further test the hypothesis that retinal remodeling, which leads to fovea formation and photoreceptor cell packing, is related to asymmetric changes in eye shape. ResultsAnole retina development follows the conventional spatiotemporal patterning observed in most vertebrates, where retina neurogenesis begins within the central retina, progresses throughout the temporal retina, and concludes in the nasal retina. One exception to this general rule is that areas that give rise to the fovea undergo retina differentiation prior to the rest of the retina. We find that retina thickness changes dynamically during periods of ocular elongation and retraction. During periods of ocular elongation, the retina thins, while during retraction it becomes thicker. Ganglion cell layer mounding is also observed in the temporal fovea region just prior to pit formation. ConclusionsAnole retina development parallels that of humans, including the onset and progression of retinal neurogenesis followed by changes in ocular shape and retinal remodeling that leads to pit formation in the retina. We propose that anoles are an excellent model system for fovea development research. Key FindingsO_LIRetina mounding that occurs in foveal areas prior to retinal differentiation progressively disappear as foveal regions of the eye elongate. C_LIO_LIThe central and temporal foveal areas undergo retina differentiation before the rest of the retina. C_LIO_LIGCL mounding prior to pit formation occurs in the area of the temporal fovea but not the central fovea. C_LIO_LIWhen the eye is experiencing ocular retraction, photoreceptor cell packing, and pit formation are observed within foveal regions. C_LI

developmental biology↗

bioRxiv2021

Chromosome-scale genome assembly of the brown anole (Anolis sagrei), a model species for evolution and ecology

Rapid technological improvements are democratizing access to high quality, chromosome-scale genome assemblies. No longer the domain of only the most highly studied model organisms, now non-traditional and emerging model species can be genome-enabled using a combination of sequencing technologies and assembly software. Consequently, old ideas built on sparse sampling across the tree of life have recently been amended in the face of genomic data drawn from a growing number of high-quality reference genomes. Arguably the most valuable are those long-studied species for which much is already known about their biology; what many term emerging model species. Here, we report a new, highly complete chromosome-scale genome assembly for the brown anole, Anolis sagrei - a lizard species widely studied across a variety of disciplines and for which a high-quality reference genome was long overdue.

genomics↗

bioRxiv2021

Anterior eye development in the brown anole, Anolis sagrei

BackgroundAnterior eye development has been explored in different vertebrate species ranging from fish to mammals. However, missing from this diverse group is a representative of reptiles. A promising candidate to fill this void is the brown anole, Anolis sagrei, which is easily raised in the laboratory and for which genome editing techniques exist. Here we provide a detailed histological analysis of the development of the anterior structures of the eye in A. sagrei, which include the cornea, iris, ciliary body, lens, trabecular meshwork, and sclera ossicles. ResultsDevelopment of the anterior segment in Anoles proceeds as for other vertebrates with the lens forming first followed by the cornea, then the iris, ciliary body, trabecular meshwork, and sclera ossicles. The onset of these latter structures occurs first temporally than nasally. Unlike the eyes of mammals and birds, anoles possess a remarkably thin cornea, flat ciliary body, and a trabecular meshwork that lacks an obvious Schlemms canal. ConclusionsThis study highlights several features present in anoles and represents an important step towards understanding reptile eye development. Key FindingsO_LIThe anole cornea epithelium is thin, composed mainly of a single basal cell layer. C_LIO_LIThe ciliary body lacks a ciliary process. C_LIO_LIIris and ciliary body formation occur in a spatiotemporal fashion, developing first temporally then nasally. C_LIO_LIThe anole trabecular meshwork is composed of a spongiform tissue and lacks a Schlemms canal. C_LI

developmental biology↗

bioRxiv2021

Ocular elongation and retraction in foveated reptiles

BackgroundPronounced asymmetric changes in ocular globe size during eye development have been observed in a number of species ranging from humans to lizards. In contrast, largely symmetric changes in globe size have been described for other species like rodents. We propose that asymmetric changes in the three-dimensional structure of the developing eye correlate with the types of retinal remodeling needed to produce areas of high photoreceptor density. To test this idea, we systematically examined three-dimensional aspects of globe size as a function of eye development in the bifoveated brown anole, Anolis sagrei. ResultsDuring embryonic development, the anole eye undergoes dynamic changes in ocular shape. Initially spherical, the eye elongates in the presumptive foveal regions of the retina and then proceeds through a period of retraction that returns the eye to its spherical shape. During this period of retraction, pit formation and photoreceptor cell packing are observed. We found a similar pattern of elongation and retraction associated with the single fovea of the veiled chameleon, Chamaeleo calyptratus. ConclusionsThese results, together with those reported for other foveated species, support the idea that areas of high photoreceptor packing occur in regions where the ocular globe asymmetrically elongates and retracts during development. Key FindingsO_LIThe eyes of the brown anole, Anolis sagrei, and veiled chameleon, Chamaeleo calyptratus undergo dynamic asymmetrical changes in ocular shape during development. C_LIO_LIIn both species, asymmetric elongation and retraction of the ocular globe is associated with fovea morphogenesis. C_LIO_LIPit formation and photoreceptor cell packing in the foveal area occur when the corresponding region of the ocular globe is retracting relative to adjacent regions. C_LI

developmental biology↗

bioRxiv2019

Pigeon foot feathering reveals conserved limb identity networks

The tetrapod limb is a stunning example of evolutionary diversity, with dramatic variation not only among distantly related species, but also between the serially homologous forelimbs (FLs) and hindlimbs (HLs) within species. Despite this variation, highly conserved genetic and developmental programs underlie limb development and identity in all tetrapods, raising the question of how limb diversification is generated from a conserved toolkit. In some breeds of domestic pigeon, shifts in the expression of two conserved limb identity transcription factors, PITX1 and TBX5, are associated with the formation of feathered HLs with partial FL identity. To determine how modulation of PITX1 and TBX5 expression affects downstream gene expression, we compared the transcriptomes of embryonic limb buds from pigeons with scaled and feathered HLs. We identified a set of differentially expressed genes enriched for genes encoding transcription factors, extracellular matrix proteins, and components of developmental signaling pathways with important roles in limb development. A subset of the genes that distinguish scaled and feathered HLs are also differentially expressed between FL and scaled HL buds in pigeons, pinpointing a set of gene expression changes downstream of PITX1 and TBX5 in the partial transformation from HL to FL identity. We extended our analyses by comparing pigeon limb bud transcriptomes to chicken, anole lizard, and mammalian datasets to identify deeply conserved PITX1- and TBX5-regulated components of the limb identity program. Our analyses reveal a suite of predominantly low-level gene expression changes that are conserved across amniotes to regulate the identity of morphologically distinct limbs.\n\nSummary statementIn feather-footed pigeons, mutant alleles of PITX1 and TBX5 drive the partial redeployment of an evolutionarily conserved forelimb genetic program in the hindlimb.

developmental biology↗

bioRxiv2019

CRISPR-Cas9 Gene Editing in Lizards Through Microinjection of Unfertilized Oocytes

CRISPR-Cas9 mediated gene editing has enabled the direct manipulation of gene function in many species. However, the reproductive biology of reptiles presents unique barriers for the use of this technology, and there are currently no reptiles with effective methods for targeted mutagenesis. Here we present a new approach that enables the efficient production of CRISPR-Cas9 induced mutations in Anolis lizards, an important model for studies of reptile evolution and development.

genetics↗