Large scale –omics projects have provided valuable knowledge on the molecular mechanisms occurring in cells and tissues of human and other related model organisms, delivering a wealth of information on the diversity of cellular processes, and in particular gene regulation. Yet, these projects have also uncovered the immense complexity of the animal cell, its potential to differentiate into various tissues, or spiral out of control by forming cancer, opening up even more profound questions on diverse fronts of the life sciences: How does tissue-specific cell differentiation work? How conserved are gene regulation mechanisms throughout animal kingdom? Where does complexity of differentiation originate from? And, finally, what is the origin of multicellularity?

Basal metazoans hold key answers to these questions: we propose to tackle them by investigating the non-model organisms at the very base of the animal evolutionary tree – the simplest multicellular animals – sponges (Porifera). Sponges, are probably the earliest branching and the oldest extant animals. They hold remarkable properties that make them an excellent subject for studying developmental gene regulation: a handful of major cell types, no true tissues and organs, and no germ layers, but at the same time they are rich in early developmental genes associated to cancer and possessing totipotent cells. With the aid of comparative genomics and transcriptomics, we wish to explore developmental and cell differentiation processes that we believe are ancestral to all multicellular animals. With a strong collaboration network, expert base in metazoan biology, expertise in genomics and computational biology of complex systems, and a preliminary set of interesting results, we propose to answer the above questions through a truly interdisciplinary approach and elucidate some of the crucial open questions in genomics and evolution of the entire animal kingdom.

Section A:

All animals inhabiting the present-day Earth are characterized by a common motif – they contain eukaryotic cells of different types, mostly grouped into tissues and organs and specialized for performing specific functions to maintain and propagate the lineage. Different cell types in one organism share an identical genetic blueprint, contained in every nucleus of each cell. The difference between cells, and subsequently tissues, originates from the processes of cell differentiation, during the development of an organism. During the developmental stages cells divide from a single zygotic cell, and characterised by a poorly understood process, undifferentiated (stem) cells activate gene regulation mechanisms that will eventually mature them into a specific, differentiated cell. Normally, this process is irreversible and cells remain in their differentiated form until their death or inactivation. Processes of developmental gene regulation as part of differentiation have been the focus of scientific research in “classical” vertebrate, insect and nematode model organisms and have revealed key genes responsible for differentiation checkpoints and cell specialization from pluripotency to a tissue. However, at the level of a complete, cellular system, the mechanistic insight into the processes governing cell differentiation are not yet understood, partly owing to the complexity of the experimental models, given the fact that vertebrate organisms contain several hundred distinct and highly specialised cell types, each undergoing a complex transformation that involves a large number of genes and an intricate regulation cascade. With such complexity of processes in vertebrate genomes, we pose the question whether it would be possible to study cell differentiation and gene regulation processes in a less-complex model with fewer cell types, especially in the context of inverse relationship between the number of cell types and their specialization in an organism. We argue that in order to truly understand the very basic processes that govern multicellular development and differentiation, we need to descend deeper towards the base of multicellular life and start exploring the mechanisms that are rooted in the last common metazoan ancestor. With this rationale, we propose the genomics study of development processes in a model that is pivotally placed at the very base of multicellular tree of life – sponges (Porifera).

Section B:
Objectives and specific aims of the proposed research

Our objective in this proposal is to systematically explore, using the -omics methodology, the base of the metazoan lineage. Only by truly understanding the processes that govern multicellularity will we be able to derive some fundamental knowledge on differentiation and development. We propose to a) collect a substantial amount of genomic information from at least three sponge classes – demosponges, calcareous sponges and hexactinellids; b) to study processes of sponge differentiation and discover the function of genes involved using transcriptomics, and c) to ellucidate the possible interaction of sponges and their endosymbionts through analyzing the prokaryote sequences obtained through high- throughput sequencing of sponge genomes.

Our specific aims are in the domain of systems biology and have a strong interdisciplinary character, as they require complementary skills in basal metazoan biology, modern –omics approaches and next generation sequencing and genome assembly, as well as the use of computational biology methods for data integration and knowledge extraction. We have therefore built a strong supporting base of collaborators who have already committed resources towards elucidating basal metazoan evolution and development, and will provide a synergistic effect to this project, ensuring its success.

Significance of the proposed project

This project presents a high-gain systems biology approach towards elucidating basal metazoan evolution, development and differentiation. If successful, it will help us answer many fundamental questions in multicellular life organization of animals, and help us define the landscape of early metazoan differentiation genes and pathways, responsible for driving the cell on its path to specialization and reverting them back to pluripotency. It will help us explain the function of genes, many of which are known only for their association with high level metabolic and sensing processes, but their primary cellular functions in simple systems such as sponges remain completely unresolved. Even the sheer processed and annotated genomic data on basal metazoan species in itself presents an indispensable contribution of this project towards resolving some of the long-standing evolutionary disputes on deep metazoan phylogeny.

Team Members and Research Group

Name Surname Academic title Institution Country
Kristian Vlahoviček (PI) PhD University of Zagreb, Faculty of Science Croatia
Helena Ćetković PhD Ruđer Bošković Institute Croatia
Matija Harcet PhD Ruđer Bošković Institute Croatia
Andreja Mikoč PhD Ruđer Bošković Institute Croatia
Rosa Karlić PhD University of Zagreb, Faculty of Science Croatia
Helena Bilandžija PhD Ruđer Bošković Institute Croatia
Mirna Imšek B.Sc. Ruđer Bošković Institute Croatia
Dragutin Perina PhD Ruđer Bošković Institute Croatia
Ivan Franulović B.Sc. University of Zagreb, Faculty of Science Croatia