Role of dHAND in the anterior-posterior polarization of the limb bud: implications for the Sonic hedgehog pathway

Abstract

dHAND is a basic helix-loop-helix (bHLH) transcription factor essential for cardiovascular development. Here we analyze its pattern of expression and functional role during chick limb development. dHAND expression was observed in the lateral plate mesoderm prior to emergence of the limb buds. Coincident with limb initiation, expression of dHAND became restricted to the posterior half of the limb bud. Experimental procedures that caused mirror-image duplications of the limb resulted in mirror-image duplications of the pattern of dHAND expression along the anterior-posterior axis. Retroviral overexpression of dHAND in the limb bud produced preaxial polydactyly, corresponding to mild polarizing activity at the anterior border. At the molecular level, misexpression of dHAND caused ectopic activation of members of the Sonic hedgehog (Shh) pathway, including Gli and Patched, in the anterior limb bud. A subset of infected embryos displayed ectopic anterior activation of Shh. Other factors implicated in anterior-posterior polarization of the bud such as the most 5′ Hoxd genes and Bmp2 were also ectopically activated at the anterior border. Our results indicate a role for dHAND in the establishment of anterior-posterior polarization of the limb bud.

A human embryonic limb cell atlas resolved in space and time

“…This included several key genes known to govern the proximal identity, including MEIS 1 & 2, PBX 1 and IRX 3 32–35 , as well as genes regulating limb outgrowth and distal morphogenesis such as WNT5A , GREM 1, ETV 4 and SALL 1 36–39 . Similarly, classical mammalian anterior-posterior (AP) genes were captured, including HAND 1, PAX 9, ALX 4 and ZIC 3 (anterior) and HAND 2, SHH and PTCH 1 and GLI 1 (posterior) 26,40–46 .…”

“…This included several key genes known to govern the proximal identity, including MEIS1 & 2, PBX1 and IRX3 [32][33][34][35] , as well as genes regulating limb outgrowth and distal morphogenesis such as WNT5A, GREM1, ETV4 and SALL1 [36][37][38][39] . Similarly, classical mammalian anterior-posterior (AP) genes were captured, including HAND1, PAX9, ALX4 and ZIC3 (anterior) and HAND2, SHH and PTCH1 and GLI1 (posterior) 26,[40][41][42][43][44][45][46] . The homeobox (HOX) genes are a group of 39 genes split into four groups termed "clusters", each of which is located on a separate chromosome.…”

(Non)Parallel developmental mechanisms in vertebrate appendage reduction and loss

“…The proximal portion of the metapterygium (dark blue) likely forms the stylopod, while the more distal elements (light blue) were likely elaborated into the distal limb structures (Ahn & Ho, 2008;Don et al, 2013;Freitas et al, 2007;Hawkins et al, 2021) F I G U R E 2 A simplified gene regulatory network implicated in vertebrate appendage development. Genes symbols coded in magenta are unique to the hindlimb, while those in blue are unique to the forelimb (Butterfield et al, 2009;Charité et al, 2000;Delgado et al, 2021;Delgado & Torres, 2015;Fernandez-Teran et al, 2000;Hockman et al, 2008;Jin et al, 2019;Lafage-Proust, 2015;McQueen & Towers, 2020;Minguillon et al, 2012;Ng et al, 2002;Nishimoto et al, 2015;Tanaka et al, 2005;te Welscher, Fernandez-Teran, et al, 2002;Xu & Wellik, 2011;Zúñiga, 2015) and provide a good system for the study of genetic parallelism of appendage reduction and loss (Bolnick et al, 2018).…”

“…Sirenians (manatees and dugongs) and cetaceans (dolphins, porpoises, and whales) are aquatic mammal lineages that have independently evolved hindlimb loss and pelvic reduction (Adam, 2009 ; Senter & Moch, 2015 ; Springer et al, 2004 ; Thewissen et al, 2001 , 2006 ). In the spotted dolphin ( Stenella attenuatus ), HAND2, an activator of Shh , is absent from the embryonic hindlimb bud (Charité et al, 2000 ; Fernandez‐Teran et al, 2000 ; Galli et al, 2010 ; Ros et al, 2003 ; Thewissen et al, 2006 ). This prevents Shh initiation which in turn diminishes Fgf8 expression (Figure 6 ) (Ros et al, 2003 ; Thewissen et al, 2006 ).…”

“… A simplified gene regulatory network implicated in vertebrate appendage development. Genes symbols coded in magenta are unique to the hindlimb, while those in blue are unique to the forelimb (Butterfield et al, 2009 ; Charité et al, 2000 ; Delgado et al, 2021 ; Delgado & Torres, 2015 ; Fernandez‐Teran et al, 2000 ; Hockman et al, 2008 ; Jin et al, 2019 ; Lafage‐Proust, 2015 ; McQueen & Towers, 2020 ; Minguillon et al, 2012 ; Ng et al, 2002 ; Nishimoto et al, 2015 ; Tanaka et al, 2005 ; te Welscher, Fernandez‐Teran, et al, 2002 ; Xu & Wellik, 2011 ; Zúñiga, 2015 ) …”