TA PROTEIN INSERTION

Uncovering the Mammalian Targeting Machinery for Tail Anchored Proteins

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 Obiettivo del progetto (Objective)

'Activation of the BCL2 oncogene underlies a wide variety of malignant transformations. BCL2 must be present on the membranes of both the mitochondria and the endoplasmic reticulum (ER) to carry out its role in protecting cells from cell death (apoptosis). To enable its anchoring to intra cellular membranes, BCL2 has a C-terminal hydrophobic tail in a unique topology shared by members of a large family of Tail Anchored (TA) proteins. In mammals hundreds of such TA proteins exist that serve diverse and essential functions on every intracellular organelle. Despite the great importance of these proteins for cellular function, the pathway for enabling their anchoring to membranes has remained unknown for many years. We have recently discovered the protein machinery required for insertion of TA proteins into ER membranes in an effective and membrane specific manner in yeast. However, the mammalian pathway is not fully characterized. Moreover, the proteins required for insertion of TA proteins into mitochondrial membranes in both yeast and mammals are completely unknown. This proposal aims at uncovering and characterizing the machinery responsible for inserting BCL2 and other TA proteins into membranes in mammals. We will take two approaches – in the first, the homologous machinery to the GET complex in mammalian cells will be explored and its affect on BCL2 localization will be studied. In parallel, we will screen for components of the cellular machinery required for insertion of TA proteins into the mitochondria using high-throughput microscopy based silencing screens in mammalian cells. The discovery of the molecular machinery guiding the sub-cellular fate of the dual-localized BCL2 oncogene, can serve as a tool for better understanding BCL2 functions and for controlling its anti-apoptotic activity. More importantly, characterizing the basic machinery required for TA protein insertion will shed light on a fundamental and little explored part of cell biology.'

Introduzione (Teaser)

Loss of optimal mitochondrial activity is implicated in a growing number of human diseases and in aging. A European study investigated the mechanism of mitochondrial protein insertion into the membrane and assessed its response under stress.

Descrizione progetto (Article)

Mitochondria are the sites of cellular energy production.

They employ specialised enzymes such as tail-anchored (TA) proteins to ensure good cell metabolism and cellular function.

Such TA proteins are usually translated in the cytosol, the cytoplasmic matrix inside any cell.

They are then inserted post-translationally into their corresponding membranes through the use of appropriate signalling pathways in the endoplasmic reticulum (ER), the cellular post office.A plethora of TA proteins exist of diverse yet essential function but the mammalian mechanism of protein insertion into membranes has not been elucidated.

The EU-funded ?Uncovering the Mammalian Targeting Machinery for Tail Anchored Proteins? (TA PROTEIN INSERTION) proposal aimed to uncover and characterise this mechanism.

Researchers focused on BCL2, an oncogene whose function is to prevent cell death.Scientists performed microscopy analysis of fluorescently tagged proteins in yeast to determine co-localisation with and dependency on other proteins.

Spf1, an ER ATPase was shown to affect mitochondrial membrane TA protein insertion by modulating lipid and protein composition.

As a result, the inherent differences in membrane composition were found to be the main determinant of determining TA protein integration specificity.

Important insight was also generated on how mitochondrial TA proteins change under different stress conditions.Apart from enhancing fundamental knowledge on the mechanisms of localisation and specificity of TA protein insertion, the project findings hold translational impact as well.

Given the involvement of BCL2 in cancer, the study outcomes should help uncover new molecular aspects of cancer, and provide new options for diagnosis and therapy.