Project Leader

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Dr. Patrick Giavalisco
Phone:+49 331 567-8623

Department Willmitzer

Experimental Systems Biology

The scientific interest of Dr. Patrick Giavalisco's research group lies in the analysis and identification of single molecules, signaling modules or entire pathways involved in the regulation of growth- and developmental processes. For this purpose we make use of classical molecular biology and genetics approaches in combination with targeted and untargeted high resolution mass spectrometry-based metabolomics and proteomics.

The aim of our approach is to generate unbiased, complex molecular patterns of proteins, metabolites and mRNA transcripts from either genetically altered or pharmacologically treated cells, including the plant model organism Arabidopsis thaliana, but also other biological systems like the green algae Chlamydomonas reinhardtii, the yeast Saccharomyces cerevisiae, the fruit fly Drosophila melanogaster, and the mold worm Caenorhabditis elegans. The measured complex and highly informative molecular patterns will then, in a next step, be further analyzed using supervised- and unsupervised statistical methods to extract and annotate significantly changing compounds, hopefully enabling us to determine relevant novel signals regulating physiology and metabolism of the organism. Especially the use of several different fully sequenced model organisms, which will be challenged with identical treatments, will be extremely valuable, since this could allow us to determine common and contrasting molecular responses, which can be further integrated into the frame of organism-wide or genome-scale molecular models.

The main biological focus of our research group lies in the elucidation of the functional implications of the Target of Rapamycin (TOR)-pathway, which has been shown to integrate environmental and nutritional information into molecular growth decisions, including general regulation of protein synthesis and metabolism. Here we are especially interested in direct but also the delayed influence of TOR-perturbations on the regulation of the central metabolism and how these regulatory functions are executed through protein- but also small molecule-based signaling cascades.

Cell proliferation and cell growth are two central aspects of every living organism. The control of these processes involves a number of complex signal transduction pathways, which as a consequence of their importance have to be tightly regulated. One of the central regulatory pathways is controlled by the Target of Rapamycin (TOR).

The Target of Rapamycin pathway in Photoautotrophic organisms

Cell proliferation and cell growth are two central aspects of every living organism. The control of these processes involves a number of complex signal transduction pathways, which as a consequence of their importance have to be tightly regulated. One of the central regulatory pathways is controlled by the Target of Rapamycin (TOR).

[more]
Optimized analysis of selected metabolites or classes of metabolites like polar amino acids or more unipolar triacylglycerols (TAG), requires specific extraction procedures in combination with specific analytical instrumentation. However, the most efficient, specific extraction protocols unfortunately often only cover a very limited number of compounds.

Analyzing proteins and metabolites: All-In-One Extraction

Optimized analysis of selected metabolites or classes of metabolites like polar amino acids or more unipolar triacylglycerols (TAG), requires specific extraction procedures in combination with specific analytical instrumentation. However, the most efficient, specific extraction protocols unfortunately often only cover a very limited number of compounds. [more]
Mass spectrometry-based analysis of metabolites and proteins has become more and more advanced in recent years. The development of this field has been driven by the continuous improvements of mass spectrometers, chromatographic separation techniques, software and the increasing quality and accessibility of public databases.

MS-based Metabolomics and Proteomics

Mass spectrometry-based analysis of metabolites and proteins has become more and more advanced in recent years. The development of this field has been driven by the continuous improvements of mass spectrometers, chromatographic separation techniques, software and the increasing quality and accessibility of public databases.

[more]
The most common approach for systematic lipid profiling is the so called shotgun lipidomic approach, which was conceptually developed more than 15 years ago. In this method, lipids are directly infused into a mass spectrometer without any further separation.

UPLC-based lipid profiling

The most common approach for systematic lipid profiling is the so called shotgun lipidomic approach, which was conceptually developed more than 15 years ago. In this method, lipids are directly infused into a mass spectrometer without any further separation. [more]
Contrary to the analysis of lipids, which show a very systematic behaviour, the analysis of the highly complex secondary metabolites from plants, offers a number of challenges. Even though many secondary metabolites are grouped into distinct classes, only few can be validated on their class-specific and systematic retention time/m/z dependencies.

UPLC-based profiling of secondary metabolites

Contrary to the analysis of lipids, which show a very systematic behaviour, the analysis of the highly complex secondary metabolites from plants, offers a number of challenges. Even though many secondary metabolites are grouped into distinct classes, only few can be validated on their class-specific and systematic retention time/m/z dependencies.

[more]
The annotation of metabolites in untargeted metabolomic analysis can be extremely tedious and often produces ambiguous, or even no results. As we and others have shown previously isotope-labelling approaches in combination with complex database searches allow not only for the bona fide distinction of biological from contaminating background peaks, but also increase the reliability of the elemental composition annotation by decreasing the false discovery rate.

Isotope labeling as a method to annotate and quantify metabolites and proteins

The annotation of metabolites in untargeted metabolomic analysis can be extremely tedious and often produces ambiguous, or even no results. As we and others have shown previously isotope-labelling approaches in combination with complex database searches allow not only for the bona fide distinction of biological from contaminating background peaks, but also increase the reliability of the elemental composition annotation by decreasing the false discovery rate. [more]
Based on the fact that the masses measured in the mass spectrometer are almost directly connected to the elemental composition of the measured analyte, we conceptualised the GoBioSpace (Golm Biochemical Space) database as a simple, repository of annotated elemental compositions, which can be directly searched with all kind of mass spectrometric data.

GoBioSpace: A database search tool for metabolite analysis

Based on the fact that the masses measured in the mass spectrometer are almost directly connected to the elemental composition of the measured analyte, we conceptualised the GoBioSpace (Golm Biochemical Space) database as a simple, repository of annotated elemental compositions, which can be directly searched with all kind of mass spectrometric data. [more]
While major improvements have been made in developing methods to analyze both primary and secondary metabolites, there are still major gaps in our understanding of the plant metabolome as a total. This is especially true with regards to the subcellular localization of metabolites and their exchange between subcellular compartments.

Subcellular distributions of metabolites and proteins within the cell: Non-aqueous fractionation (NAF)

While major improvements have been made in developing methods to analyze both primary and secondary metabolites, there are still major gaps in our understanding of the plant metabolome as a total. This is especially true with regards to the subcellular localization of metabolites and their exchange between subcellular compartments. [more]
 
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