The heart-specific proteome

The heart transcriptome

Transcriptome analysis of the heart can be visualized with regard to specificity and distribution of transcribed mRNA molecules (Figure 1). Specificity illustrates the number of genes with elevated or non-elevated expression in the heart compared to other tissues. Elevated expression includes three subcategory types of elevated expression:

• Tissue enriched: At least four-fold higher mRNA level in heart compared to any other tissues.
• Group enriched: At least four-fold higher average mRNA level in a group of 2-5 tissues compared to any other tissue.
• Tissue enhanced: At least four-fold higher mRNA level in heart compared to the average level in all other tissues.

Distribution, on the other hand, visualizes how many genes that have, or do not have, detectable levels (NX≥1) of transcribed mRNA molecules in the heart compared to other tissues. As evident in Table 1, all genes elevated in heart are categorized as:

• Detected in single: Detected in a single tissue
• Detected in some: Detected in more than one but less than one third of tissues
• Detected in many: Detected in at least a third but not all tissues
• Detected in all: Detected in all tissues

Figure 1. (A) The distribution of all genes across the five categories based on transcript specificity in heart as well as in all other tissues. (B) The distribution of all genes across the six categories, based on transcript detection (NX≥1) in heart as well as in all other tissues.

As shown in Figure 1, 387 genes show some level of elevated expression in the heart compared to other tissues. The three categories of genes with elevated expression in heart compared to other organs are shown in Table 1. In Table 2, the 12 genes with the highest enrichment in heart are defined.

Table 1. Number of genes in the subdivided categories of elevated expression in heart.

Table 2. The 12 genes with the highest level of enriched expression in heart. "Tissue distribution" describes the transcript detection (NX≥1) in heart as well as in all other tissues. "mRNA (tissue)" shows the transcript level in heart as NX values. "Tissue specificity score (TS)" corresponds to the fold-change between the expression level in heart and the tissue with second highest expression level.

Protein expression of genes elevated in heart

In-depth analysis of the elevated genes in heart using antibody-based protein profiling allowed us to visualize the expression patterns of these proteins in different functional compartments including proteins related to contraction, homeostasis or selectively expressed in intercalated discs.

Proteins specifically expressed in the heart related to contraction

To allow for the continuously beating and the long contraction period, the heart muscle is different from skeletal muscle. As a result, several proteins related to contraction are only expressed in the heart. The primary structural proteins in the heart myocytes related to contraction are myosin and actin filaments, forming a striated pattern that can be observed in electron microscopy. Another protein family related to muscular contraction is the troponin family, regulating the binding of myosin to actin via conformational changes dependent on the calcium ion concentration in the cells. Examples of members of the myosin, actin and troponin families solely expressed in heart muscle include MYH7, ACTC1 and TNNI3.

MYH7
ACTC1
TNNI3

Proteins specifically expressed in the heart related to homeostasis

In order to retain balanced levels of various substances in the body, the heart plays an important role in homeostasis. One such example is the atrial natriuretic peptide NPPA, which controls extracellular fluid volume and electrolyte homeostasis. Intracellular calcium homeostasis in the heart is achieved with calsequestrin (CASQ2) a calcium buffering protein, as well as phospholamban (PLN) a calcium pump inhibitor. Mutations in NPPA or CASQ2 are thought to be responsible for the development of cardiac arrhythmia.

NPPA
CASQ2
PLN

Proteins specifically expressed in intercalated discs of the heart

One feature unique to the heart muscle is the presence of intercalated discs, defined as the connections between two adjacent cardiomyocytes. Intercalated discs aid in the contraction of multiple cardiomyocytes simultaneously as a unit, which is necessary for proper heart function. Examples of three proteins distinctly expressed in intercalated discs are ATP1A3, CDH2 and PKP2, involved in functions related to plasma membrane ion exchange, cell adhesion and cell junctions, respectively.

ATP1A3
CDH2
PKP2

Gene expression shared between heart and other tissues

There are 133 group enriched genes expressed in heart. Group enriched genes are defined as genes showing a 4-fold higher average level of mRNA expression in a group of 2-5 tissues, including heart, compared to all other tissues.

In order to illustrate the relation of heart tissue to other tissue types, a network plot was generated, displaying the number of genes with shared expression between different tissue types.

Figure 2. An interactive network plot of the heart enriched and group enriched genes connected to their respective enriched tissues (grey circles). Red nodes represent the number of heart enriched genes and orange nodes represent the number of genes that are group enriched. The sizes of the red and orange nodes are related to the number of genes displayed within the node. Each node is clickable and results in a list of all enriched genes connected to the highlighted edges. The network is limited to group enriched genes in combinations of up to 3 tissues, but the resulting lists show the complete set of group enriched genes in the particular tissue.

Heart shares most group enriched expression with skeletal muscle, which is expected as both heart and skeletal muscle are composed as striated muscles that share similar features. Two examples of proteins group enriched in heart and skeletal muscle are MYH7 and LDB3. MYH7 is related to contraction and showed differential expression between slow (type I) and fast (type II) muscle fibers. LDB3 is involved in sarcomere organization and distinctly expressed in Z-discs.

MYH7 - heart muscle
MYH7 - skeletal muscle

LDB3 - heart muscle
LDB3 - skeletal muscle

Heart function

The heart is a coordinated muscle connected to the vascular system and specialized in pumping the blood through the body. The role of the cardiovascular system is to transport oxygen and nutrients to the cells, and to remove carbon dioxide and metabolic waste products from the body. The heart is made up of two atria and two ventricles. Deoxygenated blood from the body is pumped into the right atrium, passes through the tricuspid valve into the right ventricle, and then goes to the lungs where the blood is oxygenated and carbon dioxide is released. The oxygenated blood returns to the left atrium, and from there, it moves to the left ventricle, through the bicuspid valve and is pumped out in the body via the aorta. The heart is hence separated into two parts, which are divided by the atrioventricular septum. The left and right sides are coordinated so that the two atria contract simultaneously, and the two ventricles contract simultaneously. This is controlled by the heart's own electrical signaling system. The sinoatrial (SA) node generates electrical impulses that set the rate of contraction. The signals from the SA node also pass through another node, the atrioventricular node, which delays the signals to ensure that the atria empty completely before the ventricles contract.

Heart histology

The heart muscle is highly vascularized and under nervous control to set the pace of heartbeats. The cardiac and skeletal muscles are both composed of striated muscle tissue that forms parallel muscle fibers. However, in contrast to skeletal muscle that consists of parallel linear fibers, the cardiac muscle cells (cardiomyocytes) are arranged in fibers that exhibit cross-striations formed by alternating segments of thick and thin protein filaments.

The major cell type in the heart is cardiomyocytes, which usually contain one or two nuclei and are rich in mitochondria. They are arranged in repeating units called sarcomeres, which contain myosin and actin proteins. In the microscope, this highly structured arrangement of sarcomeres appears as dark A-bands of thick filaments and light I-bands of thin filaments. Z-discs are located at the ends of each sarcomere, in between the I-bands, and appear as dark lines. Another typical feature of the cardiac muscle, which is different from the skeletal muscle, is the dark bands, known as intercalated discs, between myocytes where the membranes of adjacent cells are situated closely together. In addition to the muscle fibers, the myocardium also includes streaks of connective tissue and cuffs of adipose tissue surrounding the smaller vessels. The cardiac muscle tissue is vascularized through the coronary arteries that branch into smaller vessels ending in a dense network of capillaries running between the fibers.

Using light sheet microscopy and immunostaining we are able to study and visualize the complexity of the heart in detail. The video below visualizes the coronary arteries in red and cardiac nerves seen in green. Full length version of the video is found here.

The histology of human heart including detailed images and information about the different cell types can be viewed in the Protein Atlas Histology Dictionary.

Background

Here, the protein-coding genes expressed in heart are described and characterized, together with examples of immunohistochemically stained tissue sections that visualize corresponding protein expression patterns of genes with elevated expression in heart.

Transcript profiling was based on a combination of three transcriptomics datasets (HPA, GTEx and FANTOM5), corresponding to a total of 9332 samples from 113 different human normal tissue types. The final consensus normalized expression (NX) value for each tissue type was used for classification of all genes according to the tissue specific expression into two different categories, based on specificity or distribution.

Relevant links and publications

Uhlén M et al., Tissue-based map of the human proteome. Science (2015)
PubMed: 25613900 DOI: 10.1126/science.1260419

Yu NY et al., Complementing tissue characterization by integrating transcriptome profiling from the Human Protein Atlas and from the FANTOM5 consortium. Nucleic Acids Res. (2015)
PubMed: 26117540 DOI: 10.1093/nar/gkv608

Fagerberg L et al., Analysis of the human tissue-specific expression by genome-wide integration of transcriptomics and antibody-based proteomics. Mol Cell Proteomics. (2014)
PubMed: 24309898 DOI: 10.1074/mcp.M113.035600

Lindskog C et al., The human cardiac and skeletal muscle proteomes defined by transcriptomics and antibody-based profiling. BMC Genomics. (2015)
PubMed: 26109061 DOI: 10.1186/s12864-015-1686-y

Histology dictionary - the heart muscle