Regulatory RNA and the origin of multicellularity

The transition from unicellular to multicellular life is among the greatest events in evolution. But it is also an event we know very little about. We know that multicellular life has evolved from unicellular ancestors a number of times. For example, plants, animals and fungi have all evolved from unicellular organisms. But the genetic mechanisms or innovations that made this possible within these particular groups is unknown. Research on the evolution of multicellularity has mostly focused on identifying genes that can explain this transition. And primarily this has meant finding genes that solely exist in multicellular groups. For example, a set of genes involved in communication between cells was for a long time exclusively found among animals. But as more and more unicellular relatives of animals were genome sequence it was found that these genes had eolved even before the first multicellular animal. More and more of these examples are being found and today we have no good idea of which genes are required to create a multicellular organism. Therefore we have in this project looked outside of the genes, in the parts of the genome popularly referred to as junk DNA. Junk DNA (the areas of the genome that are not genes in the traditional sense) has received much attention in recent years. This is primarily due to new technology that has made it possible to study all the RNA (RNA is the active parts of the genome), and not only that which comes from traditional genes. This has lead to the discovery of a whole range of new "genes". These new genes are believed to play important roles in regulating how "normal" genes are being used. For example, they have been shown to play an essential role in embryonic development in animals. But so far it is only model organisms such as mice and zebra fish that have been systematically studied for such "regulatory RNA genes". And we don't know whether these genes arose late in animal evolution and are therefore only found in "advanced animals", or whether such regulatory genes also were present even as early as the first multicellular animal? We have in this project studied the sponge Sycon ciliatum and the comb jelly Mnemiopsis leidyi which belong to the two groups of animals believed to have evolved first. In addition, we studied the unicellular species Sphaeroforma arctica which is one of the closest relatives to animals and belong to a group of species that were on Earth before animals. The first thing we did in this project examine all the expressed RNA from the sponge and identified a set of genes that could potentially represent such new regulatory RNA genes. These belonged to the "long intergenic non-coding RNAs (lincRNAs)". In order to determine a potential function of these lincRNAs we studied their expression, or how they are used during embryonic development. We did this partly through visualizing in which cells these lincRNAs were active. In addition, we showed that each stage of the embryonic development was characterized by a unique pool of lincRNAs. Furthermore, these lincRNAs was also part of gene networks together with other previously known developmental genes. We hypothesize that lincRNAs arose very early in evolution, probably before multicellular animals, and that such regulatory RNAs are also used by the more "simple" single-celled organisms. This work has been published in Proceedings B (http://rspb.royalsocietypublishing.org/content/282/1821/20151746). Parts of the results from this study was also used in a comparative study of the expression of known developmental genes between sponges and other animals. Here we showed that sponges and other animals develop their body plan similarly and that this is likely derived from the last common ancestor of all animals. This study was mainly led by Maja Adamska at the Sars Centre in Bergen and is published in Nature Communications. To follow up the work on lincRNAs in sponges we have extended this to the comb jellies to investigate whether these results also apply to other basal animal groups. Do we find lincRNAs also in comb jellies? And are these the same as in sponges? We have also studied other types of regulatory RNAs. In Sphaeroforma we have been looking for a type of very important small RNA genes called microRNAs (miRNA). miRNA is very important to fine-tune the use of other genes, for example in humans, and miRNA malfunction has been found in some cancers. Both animals and plants have miRNAs, but it is believed that these have evolved independently and they have therefore been linked to the development of multicellularity. But our hypothesis is that miRNAs were present already in the ancestors of animals, and that these organisms used miRNAs to regulate gene expression. We have looked among the shortest RNA molecules in Sphaeroforma and have identified genes we believe may be the precursors of miRNA in animals.

One of the most important evolutionary transitions in the history of life was the evolution of multicellular animals and fungi from unicellular eukaryotes. Despite this transition has been one of the most complex and profound among eukaryotes, the underly ing genetic changes of this giant leap in evolution are very much unclear. In contrast to earlier works, which focused on changes of protein coding genes, we are in the proposed project investigating alteration of gene regulation by means of small regulat ory RNAs. In recent years we have seen an increased awareness of non-coding RNA in animals and fungi. These function as regulators of gene expression, which have been found to be important for embryonic stem cell differentiation, cell proliferation, apop tosis and metabolic regulation. Hence, small regulatory RNA is vital for the development of complex body plans. A few non-coding RNAs have also been identified in pre-metazoan and pre-fungal eukaryotes, but as only a few species have so far been investiga ted, it is still a big gap in our knowledge of the processes underlining the evolution of animal and fungal multicellularity. This project aims at investigating the role of regulatory RNA in the evolution of multicellularity by comparing the genomes and R NA repertoire in unicellular relatives of animals and fungi (i.e. Choanozoa). As these two groups evolved multicellularity independently from different unicellular ancestors, these comparisons will reveal lineage-specific changes such as gene losses, expa nsions, lateral gene transfers and functional changes. The requested project will apply a combination of bioinformatic and RNA sequencing approaches to obtain a comprehensive understanding of the diversity and evolution of regulatory RNA elements within C hoanozoa and thereby also its contribution to the evolution of multicellularity.