Expression of a hindlimb-determining factorPitx1in the forelimb of the lizardPogona vitticepsduring morphogenesis

Abstract

With over 9000 species, squamates, which include lizards and snakes, are the largest group of reptiles and second-largest order of vertebrates, spanning a vast array of appendicular skeletal morphology. As such, they provide a promising system for examining developmental and molecular processes underlying limb morphology. Using the central bearded dragon (Pogona vitticeps) as the primary study model, we examined limb morphometry throughout embryonic development and characterized the expression of three known developmental genes (GHR, Pitx1 and Shh) from early embryonic stage through to hatchling stage via reverse transcription quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry (IHC). In this study, all genes were found to be transcribed in both the forelimbs and hindlimbs of P. vitticeps. While the highest level of GHR expression occurred at the hatchling stage, Pitx1 and Shh expression was greatest earlier during embryogenesis, which coincides with the onset of the differentiation between forelimb and hindlimb length. We compared our finding of Pitx1 expression—a hindlimb-determining gene—in the forelimbs of P. vitticeps to that in a closely related Australian agamid lizard, Ctenophorus pictus, where we found Pitx1 expression to be more highly expressed in the hindlimb compared with the forelimb during early and late morphogenesis—a result consistent with that found across other tetrapods. Expression of Pitx1 in forelimbs has only rarely been documented, including via in situ hybridization in a chicken and a frog. Our findings from both RT-qPCR and IHC indicate that further research across a wider range of tetrapods is needed to more fully understand evolutionary variation in molecular processes underlying limb morphology.

Interaction between physical activity,PITX1rs647161 genetic polymorphism and colorectal cancer risk in a Korean population: a case-control study

“…Although rs647161 has an unclear function, genome-wide association studies (GWAS) have recently associated the rs647161 (5q31.1) SNP with the risk of CRC [ 14 , 15 ]. PITX1 and other members of the family are involved in embryonic development [ 16 ]. Furthermore, these genes control the differentiation and proliferation of mature tissues and can play a causal role in carcinogenesis as tumor suppressor genes [ 17 ].…”

Developmental origin of limb size variation in lizards

“…While front and hind limb identity is determined by different transcription factors, once limb identity is established, the subsequent signal cascade is the same. For example, genes important to limb development of model organisms (Pitx1, Shh, and GHR) are also transcribed and expressed by embryos and hatchlings of the lizard Pogona vitticeps (Melville et al, 2016). Because genes expressed in the flank regions where limb buds will form and during limb bud formation are known from model organisms, identification of the genes responsible for interspecific differences in relative limb size of lizards could be relatively easy (e.g., Abzhanov et al, 2004;Mallarino et al, 2011).…”

“…The outgrowth of the limb itself is regulated by a feedback loop involving two fibroblast growth factors (Crossley & Martin, 1995;Mahmood et al, 1995;Martin, 1998;Tickle, 2015). While observations on gene expression in the Melville et al (2016) study were initiated late in the formation of limb buds (stage 32), they do serve to document the relevance of mouse and chicken models to limb development in lizards. For example, genes important to limb development of model organisms (Pitx1, Shh, and GHR) are also transcribed and expressed by embryos and hatchlings of the lizard Pogona vitticeps (Melville et al, 2016).…”

“…Because genes expressed in the flank regions where limb buds will form and during limb bud formation are known from model organisms, identification of the genes responsible for interspecific differences in relative limb size of lizards could be relatively easy (e.g., Abzhanov et al, 2004;Mallarino et al, 2011). For example, genes important to limb development of model organisms (Pitx1, Shh, and GHR) are also transcribed and expressed by embryos and hatchlings of the lizard Pogona vitticeps (Melville et al, 2016). While observations on gene expression in the Melville et al (2016) study were initiated late in the formation of limb buds (stage 32), they do serve to document the relevance of mouse and chicken models to limb development in lizards.…”

“…For example, genes important to limb development of model organisms (Pitx1, Shh, and GHR) are also transcribed and expressed by embryos and hatchlings of the lizard Pogona vitticeps (Melville et al, 2016). While observations on gene expression in the Melville et al (2016) study were initiated late in the formation of limb buds (stage 32), they do serve to document the relevance of mouse and chicken models to limb development in lizards.…”