Helena Nevalainen Peter Bergquist Junior Te’o Ewa Goldys Shoba Ranganathan Marko Nykänen Debra Birch Hong Yu How to retool cellular factories A systems.

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Helena Nevalainen Peter Bergquist Junior Te’o Ewa Goldys Shoba Ranganathan Marko Nykänen Debra Birch Hong Yu How to retool cellular factories A systems biology approach into understanding secretion of gene products from a fungal cell factory

Protein secretion pathway in a fungal cell factory Trichoderma reesei Fungal protein Foreign protein Secretion block ROADBLOCK -secretes protein up to 40 g/L -industrially-exploited production organism

CLSM at MU Specificity of fluorescent markers for cell organelles: H1399 and SYTO 40 for nuclei FM4-64 for membranes DiOC6(3) for ER Stability of markers in live cells Compatibility in multiple labelling EM Limited use of living cells Does not capture the dynamics of protein secretion Protein-protein interactions can be studied in fixed cells Methods

24 h live culture FM 4-64 DiOC6(3)FM 4-64 and DIOC6(3) M 4-64, DIOC6(3) and H1399

What next? Introduce the protein of interest for secretion: -gene encoding a suitable fluorescent marker -define FRET pairs for studies of the secreted protein with proteins resident in the cell organelles (interaction at the molecular level)