Tuesday March 30


Parallel session 3: Regulation of Gene Expression at the Genome Level




Molecular pathways controlling the lifespan of the ascomycete Podospora anserina

Heinz D. Osiewacz, Diana Brust, Andrea Hamann, Karin Luce

Department of Molecular Developmental Biology & Cluster of Excellence ‘Macromolecular Complexes’. Johann Wolfgang Goethe-University. Max-von-Laue-Str. 9, 60438 Frankfurt, Germany



The ascomycete Podospora anserina is characterized by a limited lifespan. After a strain specific period of growth the growth rate of a colony decreases, the morphology of the culture changes and finally the hyphal tips die. This ‘senescence syndrome’ is under control of environmental factors and genetic traits. Recent investigations uncovered a hierarchical network of pathways influencing life and healthspan of this eukaryotic ageing model. These pathways counteract molecular damage which results from natural metabolic processes (e.g. respiration). In particular, pathways involved in the control of a functional population of mitochondria were found to play a major role. At the molecular level different mitochondrial proteases are effective. Among others, PaLON, a matrix protease was found to be important in protecting cultures against the consequences of oxidative stress.


Overexpression of PaLon leads to an increased heathspan, the period in the lifespan, in which no vital functions are impaired. The corresponding transgenic strains are more resistant against oxidative stress, are characterized by reduced protein damage and improved mitochondrial function.

Although, this and other quality control pathways are effective, P. anserina cultures finally turn to senescence. During this last step in the fungal life cycle pathways leading to programmed cell death are induced. Data about investigations intervening into these pathways will be discussed.





Frequency-modulated nucleo-cytoplasmic shuttling cycles are the basis for circadian activity and abundance rhythms of the Neurospora clock transcription factor WCC

Tobias Schafmeier and Michael Brunner

Heidelberg University Biochemistry Center, INF 328, 69120 Heidelberg Germany



In Neurospora crassa the clock transcription factor White collar complex (WCC) controls the rhythmic expression of a large number of genes. The clock protein Frequency (FRQ) regulates WCC activity in a negative feedback loop by mediating its CK-1a dependent phosphorylation. In a positive loop, FRQ-dependent phosphorylation reduces the turnover rate of the WCC, resulting in accumulation of inactive and stable WCC. Rapid degradation of active WCC is a regulatory mechanism preventing an overshoot of WCC dependent transcription. WCC undergoes rapid, sub-circadian cycles of nucleo-cytoplasmic shuttling. These are linked to cycles of FRQ dependent inactivation by phosphorylation and reactivation by PP2A dependent dephosphorylation of WCC in the cytosol. Rhythmically expressed FRQ modulates the kinetics of WCC phosphorylation and shuttling cycles in a circadian manner, producing a daily rhythm of WCC activity and abundance. Hence, phosphorylation of the WCC is the molecular basis underlying both, negative feedback of FRQ on WCC activity and positive feedback of FRQ on WCC stability.





Functional characterization of LaeA

Graeme S. Garvey1, Jonathan M. Palmer2,  Jin Woo Bok1, Alex J. La Reau1, Nancy P. Keller1,3

1Department of Medical Microbiology and Immunology, University of Wisconsin-Madison

2Department of Plant Pathology, University of Wisconsin-Madison

3Department of Bacteriology, University of Wisconsin-Madison



Here we present the initial findings of a biochemical and genetic investigation into the mechanism of LaeA, a putative methyltransferase that functions as a global regulator of secondary metabolism in Aspergillus nidulans. LaeA has been found to be part of a large nuclear velvet complex that is required for secondary metabolite production as well as light regulated morphological development. Preliminary data suggests LaeA may control secondary metabolite gene clusters through chromatin remodeling. However, there is no direct evidence linking the velvet complex to chromatin remodeling. We have initiated a study to functionally characterize LaeA. Several LaeA orthologs were recombinantly expressed in E. coli and assayed for solubility. The full length LaeA protein from A. nidulans is only soluble as a MBP fusion protein, which has proved to be uninformative for in vitro activity assays. A partial proteolysis study was performed to identify soluble domains that could be amenable to in vitro analysis.  Soluble truncation mutants were identified and have proved useful for in vitro methyltransferase activity assays. Validation of the truncated LaeA proteins was carried out through successful in vivo complementation of a ∆laeA mutant. These truncation mutants are functionally equivalent to the full-length protein by restoring sterigmatocystin (ST) biosynthesis to wild type levels.  Using the truncated LaeA protein, we have confirmed binding of S-adenosyl-L-methionine (methyl group donor) and have identified a methyltransferase activity. Each candidate protein substrate’s site of methylation is being mapped by trypsin digestion coupled with LC/MS.  The in vivo role of LaeA methylation will be evaluated with point mutants for effects on ST biosynthesis.  Our findings confirm LaeA has methyltransferase activity and provide the first functional insights into the mechanism of LaeA regulation of secondary metabolism.


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