in-vivo-interrogation-central-nervous-system-translatome-polyribosome

<em>In vivo</em> Interrogation of Central Nervous System Translatome by Polyribosome Fractionation

Multiple processes are involved in gene expression including transcription, translation and stability of mRNAs and proteins. Each of these steps are ...

German Cancer Research Center (DKFZ)

In recent years, and largely supported by the increasing use of transfection technology, much research attention has been given to protein trafficking in the Plasmodium falciparum infected red blood cell. By expression of fluorescent reporter proteins, much information has been gained on both the signals and mechanisms directing proteins to their correct sub-cellular localisation within the parasite and infected host cell. Generally however, verification of the observed fluorescent phenotype is carried out using more traditional techniques such as co-immunofluorescence, protease protection, and cell fractionation followed by Western blot. Here we apply a self-assembling split GFP (saGFP) system and show that this can be used to determine both membrane topology and compartmentalisation using transfection technology alone. As an example, we verify the topology of an ER membrane protein, hDer1-1, and of an exported parasite Hsp40 co-chaperone, PFE55. Additionally, we can demonstrate that this system has the potential to be applied to analysis of organellar proteins.

Avni Baser

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