The molecular role of RE1 silencing transcription factor in uterine diseases: is there potential for targeted therapeutic development?

Loss of REST expression and resulting contribution to uterine disease pathology possess a unique challenge as the mechanisms by which this occurs are poorly understood. Several mechanistic studies have been conducted on inhibiting REST expression in context of neuronal differentiation during fetal development, but less efforts have been invested in restoration of REST expression. In leiomyomas, for example, loss of REST is not associated with enhanced β–TrCP mediated ubiquitin-proteasomal degradation of REST [ 10 ] but is associated with a loss of the planar cell polarity protein, PRICKLE1 expression [ 14 ]. PRICKLE1 associates with REST and is involved in its nuclear localization, allowing it to function as a transcription regulator [ 15 ]. As such, stabilization of PRICKLE1 to restore REST localization to the nucleus may hold promise as a therapeutic approach to restore normal signaling within uterine tissues ( Figure 2 ). Similarly, small molecule drugs which promote proper sequestration and stabilization of REST in the absence of PRICKLE1 could also aid in localization to the nucleus and proper regulation of key transcriptional signaling for normal uterine cell biology ( Figure 2 ). Given the limitations of current therapeutic approaches for treating uterine diseases, we believe targeting REST is an important new area that warrants further investigation. It should be noted that targeting REST and/or PRICKLE1 may be associated with off target effects as neither of these proteins is uterine-specific and the impact of restored or elevated expression is unknown.

The uterus is an essential female organ for reproduction which is composed of three layers, the endometrium, myometrium, and perimetrium. The endometrium is the inner most layer and functional lining of the uterus whose major function is supporting the implanting embryo and establishment and maintenance of pregnancy. The myometrium is the middle layer of the uterus and is composed primarily of smooth muscle. It plays a crucial role in uterine contractions during menstruation and labor. The perimetrium is the outermost layer of the uterus and is composed of a thin, serous membrane layer. Its primary function is to provide a protective cover for the uterus. While both the endometrium and myometrium play pivotal roles in the reproductive process, they are also susceptible to disease. Common diseases of the endometrium include endometrial cancer [ 1 ], endometriosis [ 2 ] and adenomyosis [ 3 ], while leiomyomas [ 4 ] are the most common disease of the myometrium. While endometrial cancer is malignant and can metastasize outside of the uterine body, adenomyosis and endometriosis are benign gynecological conditions where endometrial-like glands and stroma, respectively, develop within the myometrium or outside of the uterus, within the peritoneal cavity [ 2 , 3 ]. Uterine leiomyomas (uterine fibroids) are benign smooth muscle tumors of the myometrium which manifest as primarily subserosal, intramural, or submucosal tumors [ 4 ]. Despite the heterogeneity of these diseases, they share many similarities, most notably, altered responsiveness to the female sex steroids estrogen and progesterone [ 5 - 8 ]. Endometrial cancer is an estrogen-dependent disease as unopposed estrogen is a primary risk factor for disease development. Endocrine therapy (progestins or anti-estrogens) is an early form of targeted therapy for endometrial cancer, but efficacy of monotherapy is often limited, and resistance can develop over time [ 5 ]. Similarly, endometriosis and adenomyosis are both estrogen-dependent diseases with unopposed estrogen also being a risk factor for their development [ 6 , 7 ]. Treatment options for both diseases also center on progestin and/or anti-estrogen therapy and are also associated with progesterone resistance [ 6 , 7 ]. Uterine leiomyomas are characterized by enhanced estrogen sensitivity and accordingly are treated with progestins and/or anti-estrogen therapy, much like endometrial cancer, endometriosis and adenomyosis. However, similar to these other diseases, treatment of leiomyoma patients using these approaches is not without limitations, unwanted side effects, and/or lack of efficacy. Interestingly, our research group has explored the potential involvement of the epigenetic regulator of gene expression, repressor element-1 (RE1) silencing transcription factor (REST) in modulating steroid responsiveness and/or cellular events conducive to disease progression in the pathophysiology of endometrial cancer [ 9 ] and leiomyoma [ 10 , 11 ], as well as adenomyosis and endometriosis (our unpublished observations; Figure 1 ).

REST (also known as neuron-restrictive silencer factor (NRSF) and X2 box repressor (XBR), regulates gene transcription by binding to chromatin and recruiting chromatin-modifying enzymes. REST represses the transcription of neuron-specific genes, playing an important role in the development and diseases of the nervous system [ 12 ]. REST is also expressed in non-neuronal tissues including heart [ 12 ], pancreas [ 12 ] and uterus [ 9 - 11 ] among others, and loss of REST expression in these tissues is associated with pathology. REST/NRSF is a DNA-binding protein which is responsible for silencing the transcription of most neuronal differentiation genes by binding to a 21 bp consensus sequence (RE1 binding site/neuron-restrictive silencer element or RE1/NRSE), which is present at the upstream promoter – enhancer region of these genes. Upon binding to RE1/NRSE, REST recruits coregulators including CoREST, mSin3A and histone deacetylase complex (HDAC) [ 12 ]. Interestingly, the consensus RE-1/NRSF sequence is conserved among several species including human and mouse. Within the context of uterine disease, Varghese and associates reported that widespread loss of REST protein expression in uterine leiomyoma promoted aberrant gene expression and leiomyoma pathogenesis [ 10 ]. A subsequent study by Cloud and colleagues [ 11 ] reported that in vivo, loss of REST can lead to alterations in sex steroid hormone signaling and leiomyoma development. In healthy, normal myometrial tissue, REST interacts with progesterone receptor ( Figure 1 ), while loss of REST expression in leiomyoma appears to influence this interaction with its target sequences, resulting in dysregulation of REST and progesterone target genes in leiomyoma tissue ( Figure 1 ; [ 11 ]). In vitro studies from our group in which REST expression was knocked down using siRNA/shRNA resulted in activation of estrogen signaling [ 13 ] and upregulation of known REST target genes [ 9 , 13 ] as well as augmented cell migration and invasion [ 9 ]. Collectively, these studies demonstrate loss of REST expression in human endometrial cancer, endometriosis, adenomyosis and leiomyoma tissue, and in vivo and in vitro deletion/knockdown studies strongly support a functional role for loss of REST expression in mediating a cellular phenotype characteristic of these uterine pathologies. However, targeting REST as a therapeutic option for these diseases has not been explored.