Lhx1 Is Required for Specification of the Renal Progenitor Cell Field

Abstract

In the vertebrate embryo, the kidney is derived from the intermediate mesoderm. The LIM-class homeobox transcription factor lhx1 is expressed early in the intermediate mesoderm and is one of the first genes to be expressed in the nephric mesenchyme. In this study, we investigated the role of Lhx1 in specification of the kidney field by either overexpressing or depleting lhx1 in Xenopus embryos or depleting lhx1 in an explant culture system. By overexpressing a constitutively-active form of Lhx1, we established its capacity to expand the kidney field during the specification stage of kidney organogenesis. In addition, the ability of Lhx1 to expand the kidney field diminishes as kidney organogenesis transitions to the morphogenesis stage. In a complimentary set of experiments, we determined that embryos depleted of lhx1, show an almost complete loss of the kidney field. Using an explant culture system to induce kidney tissue, we confirmed that expression of genes from both proximal and distal kidney structures is affected by the absence of lhx1. Taken together our results demonstrate an essential role for Lhx1 in driving specification of the entire kidney field from the intermediate mesoderm.

osr1 Is Required for Podocyte Development Downstream of wt1a

“…21 Interestingly, we found that osr1 knockdown completely eliminated lhx1a expression specifically in the presumptive anterior pronephric region of intermediate mesoderm in 15 high-power fields embryos ( Figure 5B; n=15/15), while lhx1a expression in the tail bud of osr1 morphants remained unaltered (data not shown). We therefore tested whether lhx1a might act downstream of osr1 specifically in podocyte development by assaying whether expression of an activated form of Lhx1a (Ldb1-Lhx1; denoted LL-CA) 22 could rescue podocyte differentiation in osr1 morphants. The Lim homeodomain of Lhx1a interacts with the LIM binding protein, Ldb, and this interaction triggers Lhx1a activation.…”

“…The Lim homeodomain of Lhx1a interacts with the LIM binding protein, Ldb, and this interaction triggers Lhx1a activation. 22 We expressed a constitutively active Ldb1-Lhx1 protein (LL-CA), which is a fusion of the Ldb1 dimerization domain with the linker, C-terminal and homeodomain of zebrafish Lhx1a. 22 High doses of LL-CA mRNA (320 pg) caused early embryonic lethality associated with failed gastrulation cell movements, consistent with the known role for Lhx1a in gastrulation.…”

“…22 We expressed a constitutively active Ldb1-Lhx1 protein (LL-CA), which is a fusion of the Ldb1 dimerization domain with the linker, C-terminal and homeodomain of zebrafish Lhx1a. 22 High doses of LL-CA mRNA (320 pg) caused early embryonic lethality associated with failed gastrulation cell movements, consistent with the known role for Lhx1a in gastrulation. 23 An optimized injection dose (100 pg) allowed for 20% survival of embryos to 3 days post fertilization (n=11/50), allowing us to test whether activated Lhx1a could restore nephrin expression in osr1 morphants.…”

Heat Shock 70-kDa Protein 5 (Hspa5) Is Essential for Pronephros Formation by Mediating Retinoic Acid Signaling

“…3, A-C). Whole mount in situ hybridization and immunostaining experiments were performed to examine the formation of the pronephros in embryos targeted on one side with 30 ng of Hspa5MO2/embryo using marker genes that include pax2 (44), lhx1 (21,22), and ␤1 subunit of Na/K-ATPase (atp1b1) (45). The marker genes pax2 (60%; 18 of 30), lhx1 (67%; 24 of 36), and atp1b1 (85%; 29 of 34) were strongly suppressed on the injected side but not on the uninjected side (Fig.…”

“…The promoter region of lhx1 contains RAREs, and its expression is directly regulated by the RA signaling pathway (15). In Xenopus embryos, overexpression of lhx1 leads to the expansion of the pronephric field, whereas depletion of lhx1 severely reduces the pronephric field during organogenesis (15,21). In Xenopus, it has been reported previously that RA can induce lhx1 expression in animal caps (23).…”