A remote and highly conserved enhancer supports amygdala specific expression of the gene encoding the anxiogenic neuropeptide substance-P

Abstract

The neuropeptide substance P (SP), encoded by the preprotachykinin-A (PPTA) gene, is expressed in the central and medial amygdaloid nucleus, where it plays a critical role in modulating fear and anxiety related behaviour. Determining the regulatory systems that support PPTA expression in the amygdala may provide important insights into the causes of depression and anxiety related disorders and will provide avenues for the development of novel therapies. In order to identify the tissue specific regulatory element responsible for supporting expression of the PPTA gene in the amygdala, we used long-range comparative genomics in combination with transgenic analysis and immunohistochemistry. By comparing human and chicken genomes, it was possible to detect and characterise a highly conserved long-range enhancer that supported tissue specific expression in SP expressing cells of the medial and central amygdaloid bodies (ECR1; 158.5 kb 5 0 of human PPTA ORF). Further bioinformatic analysis using the TRANSFAC database indicated that the ECR1 element contained multiple and highly conserved consensus binding sequences of transcription factors (TFs) such as MEIS1. The results of immunohistochemical analysis of transgenic lines were consistent with the hypothesis that the MEIS1 TF interacts with and maintains ECR1 activity in the central amygdala in vivo. The discovery of ECR1 and the in vivo functional relationship with MEIS1 inferred by our studies suggests a mechanism to the regulatory systems that control PPTA expression in the amygdala. Uncovering these mechanisms may play an important role in the future development of tissue specific therapies for the treatment of anxiety and depression.

Differential Activity by Polymorphic Variants of a Remote Enhancer that Supports Galanin Expression in the Hypothalamus and Amygdala: Implications for Obesity, Depression and Alcoholism

“…Following 4 h of fixation in 4% paraformaldehyde in phosphatebuffered saline (PFA-PBS), brain sections were then stained using 5-bromo-4-chloro-3-indolyl-b-D-galactopyranoside (X-gal) solution for 4-12 h as previously described (Nagy et al, 2003). X-gal stained tissues were then prepared for vibratome sectioning as previously described (Davidson et al, 2006b). 50-mm sections were then cut on a Vibratome series 1000.…”

Genome-wide association analyses of sleep disturbance traits identify new loci and highlight shared genetics with neuropsychiatric and metabolic traits

“…For insomnia symptoms (n=32,155 cases, 26,973 controls), significant associations were observed within MEIS1 (rs113851554T, OR [95%CI]=1.26[1.20-1.33], p =9.1×10 -19 , EAF 0.057, Fig. 2b), a homeobox gene implicated in motor neuron connectivity in Drosophila 24,25 , retinal and lens development in mouse 26 , and Substance P expression in the amygdala 27 , near TMEM132E (rs145258459C, 1.23[1.13-1.35], p =2.1×10 -8 , EAF 0.983, Fig. 2c), a gene family with roles in brain development 28 , panic/anxiety 29 and bipolar disorder 30 , suggesting a link between insomnia symptoms and an underlying broader sensitivity to anxiety and stress, and near CYCL1 (rs5922858G, OR [95%CI]=1.12[1.07-1.16], p= 1.28 ×10 -8 , EAF 0.849, Fig 2d) a locus previously associated ( p =10 -6 ) with alcohol dependence co-morbid with depressive symptoms 31 .…”

The Molecular Genetics of Restless Legs Syndrome

“…49,50 The Xenopus ortholog of MEIS1 specifies neural crest cell fate, 51 and is also essential to leukemogenesis and normal hematopoiesis 52 and vascular patterning/endothelial cell development in both mice 53 and zebra fish. 54 MEIS1 also seems to regulate substance P expression in the central amygdala, which is an important gateway by which descending forebrain influences can coordinate mood with autonomic nervous system activities 55 ; two behavioral domains commonly affected by RLS. 3 As in the case of BTBD9, the inter-relationships between MEIS1 and iron homeostasis seem complex, and are not readily incorporated into heuristic constructs that posit decrements in iron storage as a necessary intermediate trait through which risk for RLS is conferred.…”