Activation of anterior HOX genes in hind- brain development during early embryo- genesis

European Bioinformatics Institute, New York University Langone Medical Center, Ontario Institute for Cancer Research, Oregon Health and Science University. The contents of this document may be freely copied and distributed in any media, provided the authors, plus the institutions, are credited, as stated under the terms of Creative Commons Attribution 4.0 International (CC BY 4.0) License. For more information see our license. This is just an excerpt of a full-length report for this pathway. To access the complete report, please download it at the Reactome Textbook. 22/12/2022

viewed in Cunningham and Duester 2015.) The RA is generated by the ALDH1A2 (RALDH2) enzyme located in somites flanking the caudal hindbrain and degraded by CYP26 enzymes expressed initially in anterior neural ectoderm of the early gastrula and then throughout most of the hindbrain (reviewed in White and Schilling 2008). HOXA1 with PBX1,2 and MEIS2 directly activate transcription of ALDH1A2 to maintain retinoic acid synthesis in the somitic mesoderm (Vitobello et al. 2011). Differentiation of embryonal carcinoma cells and embryonic stem cells in response to retinoic acid is used to model the process of differentiation in vitro (reviewed in Soprano et al. 2007, Gudas et al. 2013).
EGR2 negatively regulates HOXB1 so that by the time rhombomeres appear, HOXB1 is restricted to r4 and HOXA1 is no longer detectable (Barrow et al. 2000). EGR2 and MAFB (Kreisler) then activate HOXA3 and HOXB3 in r5 and caudal rhombomeres. Retinoic acid activates HOXA4, HOXB4, and HOXD4 in r7, the final rhombomere. HOX proteins, in turn, activate expression of genes in combination with other factors, notably members of the TALE family of transcription factors (PBX, PREP, andMEIS, reviewed in Schulte andFrank 2014, Rezsohazy et al. 2015). HOX proteins also participate in non-transcriptional interactions (reviewed in Rezsohazy 2014). In zebrafish, Xenopus, and chicken factors such as Meis3, Fgf3, Fgf8, and vHNF regulate anterior hox genes (reviewed in Schulte and Frank 2014), however less is known about the roles of homologous factors in mammals.
Mutations in HOXA1 in humans have been observed to cause developmental abnormalities located mostly in the head and neck region (Tischfield et al. 2005, Bosley et al. 2008. A missense mutation in HOXA2 causes microtia, hearing impairment, and partially cleft palate (Alasti et al. 2008). A missense mutation in HOXB1 causes a similar phenotype to the Hoxb1 null mutation in mice: bilateral facial palsy, hearing loss, and strabismus (improper alignment of the eyes) (Webb et al. 2012). Schilling, TF., White, RJ. (2008). How degrading: Cyp26s in hindbrain development. Dev. Dyn., 237, 2775-90. ↗ Duester, G., Cunningham, TJ. (2015. Mechanisms of retinoic acid signalling and its roles in organ and limb development. Nat. Rev. Mol. Cell Biol.,16,  , recruitment of coactivators such as NCOA3, and alteration of chromatin at the HOXA1 gene to an active conformation. Similar activation of HOXA1 is also observed in vitro in human breast cancer cells (Chariot et al. 1995). In mouse and Xenopus Hoxa1 acts in a feedback loop to maintain retinoic acid synthesis by directly binding and activating the promoter of the Raldh2 gene.

Literature references
In addition to recruiting transcription coactivators, retinoic acid also appears to affect histone modifications and DNA methylation. In human embryonal carcinoma cells, KDM6A (UTX) binds the HOXA1 gene upon retinoic acid treatment and demethylates trimethylated lysine-27 of histone H3 (H3K27me3) (Lee et al. 2007). Reduced H3K27me3 is also observed at HOXA1 in lung fibroblasts (Lan et al. 2007). Experiments with mouse embryos lacking Kdm6a and Kdm6b indicate other factors also participate in demethylation of H3K27me3 (Shpargel et al. 2014). Polycomb repressive complex 2 (PRC2), which binds H3K27me3, is also lost during activation by retinoic acid (inferred from mouse cells and also observed in human embryonal carcinoma cells, , Sessa et al. 2007  In mouse embryos, expression of Hoxa1 occurs in the neural tube, adjacent mesenchyme, paraxial mesoderm, somites, and gut epithelium from rhombomere 4 to the caudal-most region of the embryo. (Rhombomeres are transiently formed segments in the neural tube that will eventually form the hind- In human carcinoma cells and primary fibroblasts, KDM6A (UTX) binds the HOXB1 gene upon retinoic acid treatment (Agger et al.  and may demethylate trimethylated lysine-27 of histone H3 (H3K27me3). Reduced H3K27me3 is also observed at HOXB1 in lung fibroblasts (Lan et al. 2007). Other demethylases may be redundant with KDM6A. Polycomb repressive complex 2 (PRC2), which binds H3K27me3, is also lost during activation . KDM6A forms complexes with the histone methyltransferase KMT2C,D (MLL2,3) which may participate in methylating histone H3 at lysine-4 (H3K4me3), an activating chromatin modification . After activation by retinoic acid HOXB1 maintains its own expression by binding elements in its own promoter and activating expression (Di Rocco et al. 1997).
In mouse embryos, Hoxb1 is expressed in mesoderm and neurectoderm of primitive streak stage embryos and then becomes restricted to rhombomeres of the hindbrain. Before rhombomere formation Hoxb1 is initially expressed in the region that becomes r3-7. After rhombomere formation Hoxb1 becomes restricted to r4 and is also observed in caudal mesoderm. Hoxb1 activates expression of Egr2 (Krox20), a transcription factor that subsequently activates Hoxa2, Hoxb2, and Hoxb3 and represses Hoxb1.
As inferred from mouse embryos, HOXA1 acts synergistically with retinoic acid to activate HOXB1 (Studer et al. 1998).
Followed by: HOXB1 maintains activation of HOXB1 chromatin, HOXB1 gene is transcribed