Chamber Enriched Proteins in Cardiovascular Disease

Around the globe, cardiovascular disease is the leading cause of death. With this in mind, researchers look to cardiovascular physiology as the starting point for improving therapeutic outcomes. Unfortunately, healthy human cardiac tissues are difficult to come by, forcing researchers to rely on mouse hearts and, when available, diseased adult cardiac tissues for proteome-level analysis.

In this study, Lu et al. (2014) characterized the human cardiac chambers using human fetal atrial and ventricular samples to identify chamber-enriched and chamber-specific proteins.¹ To this end, the team collected fetal human hearts and subjected the total proteins from these samples to denaturation, reduction, alkylation and digestion before determining peptide concentration using a NanoDrop spectrophotometer (Thermo Scientific). The researchers analyzed the heart protein lysates using a Q Exactive hybrid quadrupole-Orbitrap mass spectrometer (Thermo Scientific), relying upon MaxQuant software for protein identification.

Operating in triplicate, Lu et al. identified 2,754 atrial protein groups and 2,825 ventricular protein groups. Ninety-one percent of the atrial proteins and 83% of the ventricular proteins occurred in at least two replicates. Of these, 134 atrial protein groups and 81 ventricular protein groups were chamber-enriched.

A detailed analysis of the expression levels of 3,012 unique protein groups (134 atrial-enriched, 81 ventricular-enriched and 2,797 common) highlighted several proteins previously established as cardiac chamber-enriched. These included:

Atria	Ventricles
atrial isoform of myosin light chain 2 (MYL7)	ventricular isoforms of myosin light chains (MYL2 and MYL3)
atrial natriuretic peptide (NPPA)	myosin heavy chain 7 (MYH7)
connexin 40 (GJA5)	connexin 43 (GJA1)
peptidylglycine alpha-amidating monooxygenase (PAM)

To visualize chamber-specific biological processes, the team then grouped these same 3,012 unique protein groups using Gene Ontology terms. In their report, they highlighted intracellular transport as the top process for the atrial-enriched proteins, and oxidation–reduction and muscle contraction as the top processes for ventricular-enriched proteins. For the common proteins, the major processes were localization and translation.

Next, Lu et al. used their fetal microarray data to explicate the transcriptome, revealing 8,853 total genes. They specifically looked at transcript and protein expression of the previously identified chamber-enriched proteins (134 atrial, 81 ventricular), segregating the protein groups into three groupings: high protein and transcript expression, high protein and low transcript expression, and sub-threshold transcript expression. They noted that inconsistencies between transcript and protein expression likely resulted from variations in protein/transcript stability, modifications, and transcription/translation rates.

Atria	Ventricle
high protein and transcript expression: 46 gene products	high protein and transcript expression: 45 gene products
high protein and low transcript expression: 69 gene products	high protein and low transcript expression: 25 gene products
sub-threshold transcript expression: 19 gene products	sub-threshold transcript expression: 12 gene products

Finally, the research team searched the chamber-enriched proteins against two databases: the Human Association Genomic Database and the Mouse Genome Informatics Database. Of the 134 atrial-enriched proteins, 33 had been linked with cardiovascular disease phenotypes in humans and 22 in mouse. Of the 81 ventricular-enriched proteins, 29 had been associated with cardiovascular disease phenotypes in humans and 20 in mouse.

When the team analyzed 100 randomly chosen proteins from the chamber-enriched groups, 47% had been linked with cardiovascular disease phenotypes (human or mouse), while only 33% of the total (3,012) unique proteins showed previous association with cardiovascular disease phenotypes.

Lu et al. posit that, when combined with clinical phenotype and genomic data, this proteome-level information could help researchers better understand cardiac disease. They call for further investigation to verify novel proteins in vivo and explicate how these proteins participate in cardiac disease chamber-specificity.

Reference

1. Lu, Z.Q., et al. (2014, October) “Proteomic analysis of human fetal atria and ventricle,” Journal of Proteome Research, doi: 10.1021/pr5007685 [e-pub ahead of print].

Post Author: Melissa J. Mayer. Melissa is a freelance writer who specializes in science journalism. She possesses passion for and experience in the fields of proteomics, cellular/molecular biology, microbiology, biochemistry, and immunology. Melissa is also bilingual (Spanish) and holds a teaching certificate with a biology endorsement.