Report
Teaser, summary, work performed and final results
Periodic Reporting for period 3 - RECEPIANCE (Molecular reconstitution of epigenetic centromere inheritance)
Teaser
The inheritance of genetic information through successive generations of cells or organisms is of paramount importance for the stability of life. In eukaryotic organisms (organisms in which the genetic information is confined in a specialised compartment named the nucleus)...
Summary
The inheritance of genetic information through successive generations of cells or organisms is of paramount importance for the stability of life. In eukaryotic organisms (organisms in which the genetic information is confined in a specialised compartment named the nucleus), the genetic information is stored in structures named chromosomes. During a cell\'s life, chromosomes are initially replicated and then distributed to the two daughters cells during a process known as cell division or mitosis. This process of chromosome segregation is driven by complex molecular interactions of the chromosomes with a structure named the mitotic spindle. The overarching goal of RECEPIANCE is to study the molecular basis of this process. Specifically, RECEPIANCE addresses how chromosomes build the specialised machinery that allows them to interact with the mitotic spindle to allow errorless cell division.
RECEPIANCE addresses a fundamental question on the molecular basis of genetic inheritance, which is of absolute importance for life. Understanding the molecular mechanisms of the process of chromosome segregation will also shed light on defects that lead to genetic imbalances that are typically found in human solid tumours.
The overall objective of RECEPIANCE is to perform an in vitro biochemical reconstitution of a crucial aspect of cell division. The appeal of biochemical reconstitutions is that they allow isolating and interrogating the crucial factors required to perform a given biochemical process. This, in turn, is instrumental to dissect the molecular function of such factors. In particular, RECEPIANCE aims to identify all factors required for the assembly of the structures involved in chromosome segregation, and their combination in a test tube in the presence of a source of energy that allows required biochemical work to be performed. This will not only generate a part list of the factors involved in this process, but will also allow addressing their interactions, physical relationships, and relative importance. Thus, the overall objective of RECEPIANCE is to gain a full understanding of the molecular basis of cell division.
Work performed
The RECEPIANCE program is conceived as a hierarchical organisation of progressively more complex goals, each endowed with considerable scientific interest on its own right. The ultimate goal of RECEPIANCE is the reconstitution of the mechanisms that allow the deposition on chromosomes of a protein that plays a central role in chromosome segregation. The hypothesis of RECEPIANCE is that the deposition of this protein (named CENP-A) requires a complex pre-existing structure on the chromosome (named the centromere/kinetochore). The latter is then recognised, through mechanism of macromolecular interaction, by protein machinery specifically devoted to the task of depositing new CENP-A. Since the inception of the program, the RECEPIANCE team dedicated itself to the reconstitution of this chromosome structure (the centromere/kinetochore) in vitro and to the identification of new factors required for the deposition of CENP-A. Considerable progress was achieved in both areas of research. For instance, a major intermediate achievement of RECEPIANCE was the reconstitution of a near complete centromere/kinetochore particle (Weir et al. Nature 2016) which will be used as substrate in the reaction of CENP-A deposition.
Final results
\"Studies of CENP-A and its role in the function of centromeres/kinetochores have a long tradition that dates back to the mid 1980s, when CENP-A was first discovered. Our work goes well beyond the state of the art, as it attempts the reconstitution in vitro of an extremely complex molecular reaction probably requiring 50 or more different polypeptide chains and a complex chromosome structure. This is an extreme level of ambition and we have reasons to believe that it will likely propel the centromere/kinetochore field in a new dimension, one in which all crucial molecular interactions will have been identified, characterised, and understood in molecular terms. Our work is a good representation of the concept, expressed by famous physicist Richard Feynman, that \"\"what I cannot create, I do not understand\"\". By (re)creating the reaction through reconstitution, we will finally be able to understand it in its molecular essence.\"