Researcher Discovers New Regulatory Autism Gene
WASHINGTON (July 1, 2013) – A new study by Valerie Hu, Ph.D., professor of biochemistry and molecular medicine at the George Washington University (GW) School of Medicine and Health Sciences (SMHS), reports that RORA, a novel candidate gene for autism discovered by her group in a 2010 study, regulates a large number of other genes associated with autism.
“We are focusing on this gene, in part, because this gene can act as a master regulator of other genes,” said Hu, whose study was published in the journal Molecular Autism. “Called nuclear hormone receptors, they are capable of activating or suppressing other genes in the genome. The question was which specific genes are regulated by RORA.”
Hu and co-author, Tewarit Sarachana, Ph.D., a former doctoral student in the molecular medicine doctoral program at SMHS, found that RORA encodes a protein that can regulate the expression of more than 2,500 other genes. Of these 2,500 genes, many are known to be involved in neuronal development and functions, and 426 of RORA’s gene targets are already listed in AutismKB, a database of known autism candidate genes.
To identify genes regulated by RORA, Hu and Sarachana used chromatin immunoprecipitation (ChIP) with an anti-RORA antibody followed by whole-genome promoter array (chip) analysis. This genome-wide ChIP-on-chip analysis of target genes of RORA, as well as additional methods of validation, confirmed that RORA transcriptionally regulates the genes A2BP1, CYP19A1, HSD17B10, ITPR1, NLGN1, and NTRK2, such that when RORA levels are cut in half, all six genes also go down in their expression. The expression levels of these six genes are also reduced in RORA-deficient postmortem brain tissues from individuals with autism relative to that of age-matched unaffected controls.
“We see it as a domino effect, where RORA is a particularly shaky domino,” said Hu. “If knocked over, it can also knock down a whole bunch of other genes, except that it’s not just a single chain of events. There are multiple chains of events, leading to massive disruption of gene expression in autism.”
A 2011 study by Hu’s group revealed that RORA has the potential to be under negative and positive regulation by androgen and estrogen, respectively, suggesting that RORA may also contribute to the male bias of autism spectrum disorder.
The new study, titled “Genome-wide identification of transcriptional targets of RORA reveals direct regulation of multiple genes associated with autism spectrum disorder,” is available online at http://www.molecularautism.com/content/4/1/14.
From the Full Paper – http://www.molecularautism.com/content/4/1/14.
” We have recently identified a novel autism candidate gene, retinoic acid-related (RAR) orphan receptor-alpha (RORA)  which is regulated by male and female hormones in a manner that may provide an explanation for the higher testosterone levels and, possibly, sex bias in ASD .
RORA is a ligand-dependent orphan nuclear hormone receptor that, in combination with co-regulator proteins, serves as a transcriptional regulator. Although RORA has never before been associated with ASD, our recent studies have demonstrated: reduced expression of RORA in lymphoblastoid cell lines (LCL) derived from individuals with autism ; increased methylation leading to reduced expression of RORA in the LCL from cases vs. sibling controls ; and decreased expression of RORA protein in the prefrontal cortex and the cerebellum of individuals with autism . Together, these results link these molecular changes in RORA in blood-derived peripheral cells to molecular pathology in the brain tissues of individuals with autism.
These findings are notable because studies on the Rora-deficient staggerer mouse model indicate that Rora is involved in several processes potentially relevant to autism,
protection of neurons against oxidative stress ,
suppression of inflammation , and
regulation of circadian rhythm .
Indeed, the involvement of Purkinje cells and cerebellar abnormalities as well as neuroinflammation and oxidative stress in the autistic brain has been comprehensively discussed in a consensus report on the pathological role of the cerebellum in autism .”
Further Readings of Interest
Gene expression profiling of lymphoblastoid cell lines from monozygotic twins discordant in severity of autism reveals differential regulation of neurologically relevant genes
“One of the striking results of the pathway analyses is that a relatively large number of the differentially expressed, neurologically relevant genes are linked in networks that are centered on genes involved in inflammation (see Figs. 1 and 2).
The network genes with reported neurological functions include the proteins ASS, ALOX5AP (FLAP), CD44, CHL1, DAPK1, EGR2, F13A1, FLT1, IL6ST, NAGLU, PTGS2, and ROBO1 (See Table 3).
The protein ASS regulates the rate-limiting step involved in nitric oxide (NO) production through regeneration of arginine from citrulline, a byproduct of the nitric oxide synthetase (NOS) reaction . Since NO is a major signaling molecule in the brain that has been implicated in several psychiatric disorders, including autism , the increased expression of ASS may be of potential relevance to the autistic phenotype.
Interestingly, the expression of FLT1 (VEGF receptor 1) is also regulated by inflammatory cytokines as well as by NO .
Furthermore, the fact that IL6ST (gp130) is increased in LCL from the more severely affected twin, may complement previous observations that IL-6 is the most elevated inflammatory cytokine in the middle frontal gyrus and anterior cingulate gyrus of brain autopsy tissue from autistic individuals .
While upregulation of ASS, DAPK1, FLT1, and IL6ST may be responses to inflammation, ALOX5AP (FLAP) and PTGS2 (COX-2) mediate inflammation through the production of leukotrienes  and prostaglandins .
Interestingly, 5-lipoxygenase, the target of FLAP activation, has been implicated in aging and neurodegenerative diseases , as well as other psychiatric disorders , including anxiety and depression, which are frequently co-morbid conditions of autism, while a COX-2 inhibitor, celecoxib, has been shown to have therapeutic effects in major depression , further suggesting a role for inflammatory processes in psychiatric disease.
Collectively, the potential involvement of these specific genes that are associated with neurological function and disease and their presence in pathways regulated by inflammatory mediators lend further support to the neural inflammation model for autism , which may be also manifested by immune dysfunctions commonly observed in autism .
The orphan nuclear receptor ROR alpha is a negative regulator of the inflammatory response.
Retinoid-related orphan receptor alpha (ROR alpha) (NR1F1) is a member of the nuclear receptor superfamily whose biological functions are largely unknown.
Since staggerer mice, which carry a deletion in the ROR alpha gene, suffer from immune abnormalities, we generated an adenovirus encoding ROR alpha1 to investigate its potential role in control of the inflammatory response.
We demonstrated that ROR alpha is expressed in human primary smooth-muscle cells and that ectopic expression of ROR alpha1 inhibits TNFalpha-induced IL-6, IL-8 and COX-2 expression in these cells.
ROR alpha1 negatively interferes with the NF-kappaB signalling pathway by reducing p65 translocation as demonstrated by western blotting, immunostaining and electrophoretic mobility shift assays.
This action of ROR alpha1 on NF-kappaB is associated with the induction of IkappaB alpha, the major inhibitory protein of the NF-kappaB signalling pathway, whose expression was found to be transcriptionally upregulated by ROR alpha1 via a ROR response element in the IkappaB alpha promoter.
Taken together, these data identify ROR alpha1 as a potential target in the treatment of chronic inflammatory diseases, including atherosclerosis and rheumatoid arthritis.