Associate Investigator, TIGEM
Associate Professor of Medical Genetics, Department of Biochemistry, Biophysics and General Pathology, Second University of Naples, Italy
The Role of Noncoding RNAs in Eye Function and Disease and the study of the molecular basis of eye inherited disorders
The progress of the Human Genome Project has led to a revolutionary change in the strategies used for identifying and cloning genes in the last ten years. Advanced transcriptome techniques including next generation sequencing, can be used to identify coding or non-coding genes that are possibly involved in the pathogenesis of human inherited eye disorders.
Inherited eye diseases are a common cause of visual impairment in children and young adults. The molecular causes of a significant fraction of a large variety of ocular genetic disorders recognized thus far have not been elucidated yet. In particular, we are interested in understanding the pathogenic mechanisms underlying inherited retinal disorders, including retinitis pigmentosa (RP) and Leber congenital amaurosis (LCA) and eye developmental anomalies such as microphthalmia, anophtalmia and coloboma.
We are currently focusing on elucidating the role of microRNAs in eye development and function. MicroRNAs are small, endogenous RNAs that negatively regulate gene expression post-transcriptionally by binding to target sites in the 3’ untranslated region (UTR) of messenger RNAs. We first identified seven microRNAs expressed in the eye with both differing and partially overlapping patterns, which may reflect their roles in controlling cell differentiation of the retina as well as of other ocular structures. More recently, we generated the most comprehensive atlas of microRNA expression data in the eye available to date, based on both high-throughput transcriptome analysis by microarray and RNA in situ hybridization. We are currently developing a more defined retinal miRNome/transcriptome by using next generation sequencing approaches. Finally, we demonstrated, by using variety of in vivo and in vitro approaches, that one of the eye-expressed microRNAs we identified in the latter efforts, i.e., miR-204, regulates multiple aspects of eye development in the medaka fish (Oryzias latipes). In particular, we found that the ablation of miR-204 expression resulted in an eye phenotype characterized by microphthalmia, abnormal lens formation, and altered dorsoventral (D-V) patterning of the retina, which is associated with optic fissure coloboma. Using a variety of in vivo and in vitro approaches, we were able to identify several key target mRNAs that play an important role in miR-204 function, including the Meis2 transcription factor and the Ankrd13a gene. We showed that this microRNA is able to precisely regulate the gene pathway controlled by Pax6, a master regulator of eye development. Our data provide an example of how a specific miRNA can regulate multiple events in eye formation.
To gain insight on the function of specific microRNAs it is important to identify their mRNA targets. Towards this goal, we first developed the HocTAR tool, which recognized the targets of intragenic microRNAs by combining the expression analysis of their host genes and sequence prediction softwares (http://hoctar.tigem.it). More recently, we further exploited the integration of co-expression data analysis and of target prediction and implemented the co-expression Meta-analysis of miRNA Targets (CoMeTa) tool to gain insight into the biological functions of miRNAs and to provide a comprehensive, genome-wide scale analysis of human miRNA regulatory networks.
Identification of mRNA targets is key to gaining insight of the function of specific microRNAs. Towards this goal, we also developed a novel strategy combining expression data analysis and sequence prediction softwares. The results of our prediction are publicly available at http://hoctar.tigem.it.
Relevant databases generated by our lab are as follows:
Host gene Opposite Correlated TARgets databases (http://hoctar.tigem.it)
miRNeye expression atlas (http://mirneye.tigem.it)
COmparative GEnomics MIcroRna database (http://cogemir.tigem.it)
RP Gene expression Atlas Database (http://www.tigem.it/RPexp/)
Gennarino VA, D'Angelo G, Dharmalingam G, Fernandez S, Russolillo G, Sanges R, Mutarelli M, Belcastro V, Ballabio A, Verde P, Sardiello M, Banfi S (2012). Identification of microRNA-regulated gene networks by expression analysis of target genes. Genome Res. 22:1163-1172. doi: 10.1101/gr.130435.
Conte I, Merella S, Garcia-Manteiga JM, Migliore C, Lazarevic D, Carrella S, Marco-Ferreres R, Avellino R, Davidson NP, Emmett W, Sanges R, Bockett N, Van Heel D, Meroni G, Bovolenta P, Stupka E, Banfi S (2014). The combination of transcriptomics and informatics identifies pathways targeted by miR-204 during neurogenesis and axon guidance. Nucleic Acids Res. 42(12):7793-806. doi: 10.1093/nar/gku498.
Conte I, Carrella S, Avellino R, Karali M, Marco-Ferreres R, Bovolenta P, Banfi S (2010). miR-204 is required for lens and retinal development via Meis2 targeting. Proc Natl Acad Sci USA. 107: 15491-15496. doi: 10.1073/pnas.0914785107.
Karali M, Peluso I, Gennarino VA, Bilio M, Verde R, Lago G, Dollé P, Banfi S (2010). miRNeye: a microRNA expression atlas of the mouse eye. BMC Genomics. 11: 715. doi: 10.1186/1471-2164-11-715.
Gennarino VA, Sardiello M, Avellino R, Meola N, Maselli V, Anand S, Cutillo L, Ballabio A, Banfi S (2009). MicroRNA target prediction by expression analysis of host genes. Genome Res. 19: 481-490. doi:10.1101/gr.084129.108.