Left-right function of dmrt2 genes is not conserved between zebrafish and mouse

Abstract

Background: Members of the Dmrt family, generally associated with sex determination, were shown to be involved in several other functions during embryonic development. Dmrt2 has been studied in the context of zebrafish development where, due to a duplication event, two paralog genes dmrt2a and dmrt2b are present. Both zebrafish dmrt2a/terra and dmrt2b are important to regulate left-right patterning in the lateral plate mesoderm. In addition, dmrt2a/terra is necessary for symmetric somite formation while dmrt2b regulates somite differentiation impacting on slow muscle development. One dmrt2 gene is also expressed in the mouse embryo, where it is necessary for somite differentiation but with an impact on axial skeleton development. However, nothing was known about its role during left-right patterning in the lateral plate mesoderm or in the symmetric synchronization of somite formation.

Methodology/principal findings: Using a dmrt2 mutant mouse line, we show that this gene is not involved in symmetric somite formation and does not regulate the laterality pathway that controls left-right asymmetric organ positioning. We reveal that dmrt2a/terra is present in the zebrafish laterality organ, the Kupffer's vesicle, while its homologue is excluded from the mouse equivalent structure, the node. On the basis of evolutionary sub-functionalization and neo-functionalization theories we discuss this absence of functional conservation.

Conclusions/significance: Our results show that the role of dmrt2 gene is not conserved during zebrafish and mouse embryonic development.

Figure 1. Dmrt2 is not required for symmetric somite formation in the mouse embryo.

(A–P) Expression pattern of the cyclic genes hes7 (A–E), lfng (F–J), axin2 (K–M) and sprouty2 (N–P) in E8.5 mouse embryos. (A–J) Embryos were also hybridized with the somite marker uncx4.1. (A, F, K, N) WT, (B, G, L, O) heterozygous and (C–E, H–J, M, P) dmrt2 homozygous mutant embryos show the same phase of hes1, hes7, axin2 and sprouty2 cyclic expression in the PSM. All views are dorsal with anterior to the top.

Figure 2. Dmrt2 is not required for the left-right LPM patterning in the mouse embryo.

(A–F) Embryos between E8.0 and E8.5 processed by whole mount in situ hybridization with nodal (A–C) and pitx2 (D–F) probes, respectively. (A–C) nodal expression is restricted to the left side of the node and left LPM in (A) WT, (B) heterozygous and (C) dmrt2 homozygous mutant embryos. (D–F) pitx2 expression is also restricted to the left LPM in (D) WT, (E) heterozygous and (F) dmrt2 homozygous mutant embryos. (G–L) Localization of the internal organs in newborn mice. (G) In WT, (H) heterozygous and (I) dmrt2 homozygous mutants the heart always bend to the left, the left lung always has one lobe (ll) and the right lung has four lobes (ml, crl, cl, al). (J–L) Concomitant with the normal situs of the heart and lungs, the stomach is also always placed on the correct left side of the axis. ml (middle lobe); crl (cranial lobe); cl (caudal lobe); al (accessory lobe); ll (left lobe); h (heart); s (stomach). All views are ventral with anterior to the top.

Figure 3. dmrt2/terra is expressed in the zebrafish kupfer's vesicle.

(A–F) Whole mount in situ hybridization confirming the dmrt2 conserved expression pattern in the anterior PSM and somites in mice (A–C) and zebrafish (D–F) embryos. Despite not being expressed in the mouse node (dashed circle) (A–C), dmrt2a/terra is present in the zebrafish equivalent structure, the kupffer's vesicle (arrow) (D–F). (A–C) Ventral views of mice embryos with anterior to the top. (D–F) Dorsal views of zebrafish embryos with anterior to the top. S, somite stage.