Sensing Small Changes in Protein Abundance: Stimulation of Caco‑2 Cells by Human Whey Proteins

Mass spectrometry (MS)-based proteomic
approaches have largely
facilitated our systemic understanding of cellular processes and biological
functions. Cutoffs in protein expression fold changes (FCs) are often
arbitrarily determined in MS-based quantification with no demonstrable
determination of small magnitude changes in protein expression. Therefore,
many biological insights may remain veiled due to high FC cutoffs.
Herein, we employ the intestinal epithelial cell (IEC) line Caco-2
as a model system to demonstrate the dynamicity of tandem-mass-tag
(TMT) labeling over a range of 5–40% changes in protein abundance,
with the variance controls of ±5% FC for around 95% of TMT ratios
when sampling 9–12 biological replicates. We further applied
this procedure to examine the temporal proteome of Caco-2 cells upon
exposure to human whey proteins (WP). Pathway assessments predict
subtle effects due to WP in moderating xenobiotic metabolism, promoting
proliferation and various other cellular functions in differentiating
enterocyte-like Caco-2 cells. This demonstration of a sensitive MS
approach may open up new perspectives in the system-wide exploration
of elusive or transient biological effects by facilitating scrutiny
of narrow windows of proteome abundance changes. Furthermore, we anticipate
this study will encourage more investigations of WP on infant gastrointestinal
tract development.