Secretomics

Secretomics is a subset of proteomics in which all of the secreted proteins of a cell, tissue, or organism are analyzed. Secreted proteins are involved in a variety of physiological processes, including cell signaling and matrix remodeling, but are also integral to invasion and metastasis of malignant cells. Secretomics has thus been especially important in the discovery of biomarkers for cancer.

In 2000 Tjalsma et al. coined the term ‘secretome’ in their study of the eubacterium B. subtilis. They defined the secretome as all of the secreted proteins and secretory machinery of the bacteria. Using a database of protein sequences in B. subtilis and an algorithm that looked at cleavage sites and amino-terminal signal peptides characteristic of secreted proteins they were able to predict what fraction of the proteome is secreted by the cell. In 2001 the same lab set a standard of secretomics – predictions based on amino acid sequence alone are not enough to define the secretome. They used two-dimensional gel electrophoresis and mass spectrometry to identify 82 proteins secreted by B. subtilis, only 48 of which had been predicted using the genome-based method of their previous paper. This demonstrates the need for protein verification of predicted findings.

As the complicated nature of secretory pathways was revealed – namely that there are many non-classical pathways of secretion and there are many non-secreted proteins that are a part of the classical secretory pathway – a more in-depth definition of the secretome became necessary. In 2010, Agrawal et al. suggested defining the secretome as “the global group of secreted proteins into the extracellular space by a cell, tissue, organ, or organism at any given time and conditions through known and unknown secretory mechanisms involving constitutive and regulated secretory organelles.”

In culture, cells are surrounded by contaminants. Bovine serum from cell culture media and cellular debris can contaminate the collection of secreted proteins used for analysis. Bovine contaminants present a particular challenge because the protein sequences of many bovine extracellular proteins, like fibronectin and fibulin-1, are similar to the human protein sequences. To remove these contaminants, cells can be washed with PBS or serum-free medium (SFM) before incubating in SFM and collecting secreted proteins. Care must be taken not to burst cells, releasing intracellular proteins. In addition, incubation time and conditions must be optimized so that the metabolic stress that can be induced by the lack of nutrients in SFM does not affect secretomic analysis.

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