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Long intervening noncoding RNAs (lincRNAs) in vertebrate development

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Group leader : Alena Shkumatava
Long intervening noncoding RNAs (lincRNAs) in vertebrate development

Keywords

lincRNAs, regulatory RNAs, zebrafish development, brain development, genomics

Plain english

The human genome encodes thousands of long intervening noncoding RNAs (lincRNAs) that have been called the “dark matter” of the genome because little is known about their functions. Our lab studies the roles of lincRNAs in embryonic development using zebrafish as a tool. We perturb selected lincRNAs in fish, observe how loss of a lincRNA affects embryonic development and then replace it with the human variant of the same lincRNA. Two lincRNAs that we looked at restore proper development in zebrafish, indicating that these non-coding RNAs have equivalent functions in humans and fish.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

To understand developmental functions of lincRNAs, we have identified over 550 lincRNAs in zebrafish. We previously demonstrated that two novel lincRNAs, megamind and cyrano, are required for proper embryonic development, in particular for brain morphogenesis and neurogenesis (Figure 1) (Ulitsky*, Shkumatava* et al., Cell, 2011). Remarkably, the functions of megamind and cyrano are conserved between zebrafish and mammals, despite rapid evolution that has resulted in little sequence conservation. Our goal is to determine the functions of lincRNAs in vertebrate development and identify molecular mechanisms they employ to carry out these functions. 

 

Developmental functions of lincRNAs


The majority of lincRNAs are expressed in specific regions of the developing nervous system in zebrafish embryos (Figure 2). Thus, we are particularly interested in the roles lincRNAs play in establishing cellular complexity in CNS. We are exploring lincRNA functions by manipulating their gene expression and analyzing phenotypic consequences of lincRNA perturbations in zebrafish embryos. We are focusing on lincRNAs that, like megamind and cyrano, have sequence conservation to mammals.

Figure 1: The importance of cyrano for proper embryonic development. Embryos injected with anti-sense morpholinos targeting cyrano show curly tails, small heads (top panel) and enlargement of the nasal placode (arrows; bottom panel) (Ulitsky*, Shkumatava* et al., Cell, 2011).Figure 1: The importance of cyrano for proper embryonic development. Embryos injected with anti-sense morpholinos targeting cyrano show curly tails, small heads (top panel) and enlargement of the nasal placode (arrows; bottom panel) (Ulitsky*, Shkumatava* et al., Cell, 2011).

Molecular and biochemical mechanisms of lincRNAs action

To unravel how lincRNAs regulate embryonic development, we are investigating the molecular and biochemical mechanisms of lincRNA action by determining their interaction partners and downstream targets. We use two complementary systems, zebrafish embryos and mammalian cell culture, to dissect cellular roles of lincRNAs with important biological functions, such as megamind and cyrano. The powerful combination of zebrafish genetics, molecular and cellular biology, genome-wide approaches and bioinformatics allows us to address whether lincRNAs employ common mechanisms of action similar to other noncoding RNAs such as microRNAs.

Figure 2: Tissue-specific expression of lincRNAs by in situ hybridization in zebrafish embryos (Ulitsky*, Shkumatava* et al., Cell, 2011).Figure 2: Tissue-specific expression of lincRNAs by in situ hybridization in zebrafish embryos (Ulitsky*, Shkumatava* et al., Cell, 2011).

The origin and evolution of vertebrate lincRNA genes

lincRNAs represent a rapidly evolving group of important developmental regulators: only 5.1% of zebrafish lincRNAs have detectable sequence conservation with mammalian lincRNAs. Moreover, the sequence similarity usually spans a small portion of the transcript, typically restricted to a single exon. Nevertheless, while requiring a relatively small amount of specific sequence, the function of some lincRNAs is conserved between distant vertebrate species. To better understand determinants of lincRNA functionality, our lab is investigating the origin and evolution of lincRNA.

Figure 3: Cytoplasmic localization of lincRNA cyrano in the zebrafish retina by single molecule in situ hybridization. Nuclei (blue), cyrano (red), membrane GFP (green)Figure 3: Cytoplasmic localization of lincRNA cyrano in the zebrafish retina by single molecule in situ hybridization. Nuclei (blue), cyrano (red), membrane GFP (green)

 

Key Publications

Year of publication: 2011
Ulitsky I*, Shkumatava A*, Jan C, Sive H, Bartel DP. Conserved function of lincRNAs in vertebrate embryonic development despite rapid sequence evolution.
Cell. 147(7):1537-1550.
* denotes equally contributing authors

Year of publication: 2009
Shkumatava A, Stark A, Sive H and Bartel DP. Coherent but overlapping expression of miRNAs and their targets during vertebrate development.
Genes Dev. 15;23(4):466-81.