The smooth muscle-specific proteome
Smooth muscle fibers are found throughout the body in blood vessels and hollow organs. Through their ability to apply pressure by involuntary muscle contraction, they are able to regulate essential bodily functions, such as blood pressure and bowel movement. During contraction, dense bodies are used by smooth muscle cells as anchoring points for the actin and intermediate filaments to exert force upon. Smooth muscle fibers are built up of smooth muscle cells attached to each other using gap junctions to synchronize their response to stimuli. Transcriptome analysis shows that 66% (n=13166) of all human proteins (n=20090) are expressed in the smooth muscle and 47 of these genes show an elevated expression in the smooth muscle compared to other tissue types.
The smooth muscle transcriptome
Transcriptome analysis of the smooth muscle can be visualized with regard to the specificity and distribution of transcribed mRNA molecules (Figure 1). Specificity illustrates the number of genes with elevated or non-elevated expression in the smooth muscle compared to other tissues. Elevated expression includes three subcategory types of elevated expression:
Distribution, on the other hand, visualizes how many genes have, or do not have, detectable levels (nTPM≥1) of transcribed mRNA molecules in the smooth muscle compared to other tissues. As evident in Table 1, all genes elevated in smooth muscle are categorized as:
Figure 1. (A) The distribution of all genes across the five categories based on transcript specificity in smooth muscle as well as in all other tissues. (B) The distribution of all genes across the six categories, based on transcript detection (nTPM≥1) in smooth muscle as well as in all other tissues.
As shown in Figure 1, 47 genes show some level of elevated expression in smooth muscle compared to other tissues. The three categories of genes with elevated expression in smooth muscle compared to other organs are shown in Table 1.
Table 1. The number of genes in the subdivided categories of elevated expression in smooth muscle.
Protein expression of genes elevated in smooth muscle
In-depth analysis of the elevated genes in smooth muscle using antibody-based protein profiling allowed us to visualize the expression patterns of these proteins in different functional compartments including genes involved in contraction and calcium homeostasis.
Proteins related to contraction
The primary structural proteins related to the contraction in smooth muscle cells are myosin and actin filament proteins. Another protein family related to muscular contraction is the filamin family, regulating the binding of actin to membranous glycoproteins. The typical smooth muscle actins; ACTG2 and ACTA2 are shown below.
Proteins related to calcium homeostasis
Smooth muscle contraction is dependent on the level of intracellular calcium. Smooth muscle cells, like skeletal myocytes, store calcium in the sarcoplasmic reticulum until a neuronal impulse triggers calcium influx along the smooth muscle cells. Two genes involved in the regulation of muscle contraction via calcium interaction are Caldesmon 1 (CALD1) and Caponin 1 (CNN1).
Gene expression shared between smooth muscle and other tissues
There are 7 group enriched genes expressed in smooth muscle. 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 smooth muscle, compared to all other tissues.
To illustrate the relation of smooth muscle tissue to other tissue types, a network plot was generated, displaying the number of genes with a shared expression between different tissue types.
Figure 2. An interactive network plot of the smooth muscle enriched and group enriched genes connected to their respective enriched tissues (grey circles). Red nodes represent the number of smooth muscle 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 5 tissues, but the resulting lists show the complete set of group enriched genes in the particular tissue.
As shown in figure 2, few genes elevated in smooth muscle fit the criteria for group enrichment. This is mainly attributed to the fact that smooth muscle is found in exceptionally many tissues, resulting in most of these genes being categorized as low tissue specificity genes. Smooth muscle shares elevated gene expression with many of the female tissues since these contain a great amount of smooth muscle cells as can be seen in for example PGR. Besides other tissues containing smooth muscle cells this tissue type shares genes with other muscle types like for example tongue and skeletal muscle. An example of such a gene is SYNC that is a intermediate filament protein that assists in the transfer of the mechanical forces from the muscle fibers onto the exterior.
Smooth muscle function
Smooth muscle fibers can be found throughout the body in, among other places; blood vessels, eyes and hollow organs like the bladder, uterus, intestine and stomach. The fibers can exert force on the tissue they are in or around, enabling blood vessel constriction and movement of food through the intestine. Unlike skeletal and heart muscle, smooth muscle fibers do not contain Z-discs, but instead contain dense bodies which are used as the anchoring point for the actin and intermediate filaments.
Smooth muscle histology
Smooth muscle tissue can be either of the single- or multi-unit type. The single-unit smooth muscle tissue is composed of non-striated myocytes that form parallel muscle fibers. The smooth muscle cells are non-striated since the actin and myosin are more randomly organized inside the cell. Multiple smooth muscle cells organize themselves in fibers using gap junctions. Longitudinal as well as transverse fibers can be observed in stained sections. Multi-unit smooth muscle cells can often be found surrounding ducts and small blood vessels and have typically no gap junctions.
The typical smooth muscle cells are small spindle-shaped cells of approximately 30-200 μm long with a single central nucleus.
The activity of smooth muscle is regulated via the autonomous nervous system. Axons around the smooth muscle stimulate smooth muscle activity by secreting neurotransmitters through their boutons, also called varicosities. In single-unit smooth muscle tissue, the smooth muscle cells get stimulated as a group because they are linked via their gap junctions. This way, the nerve does not need to penetrate the muscle tissue, as is the case in the intestine. In contrast, in the multi-unit smooth muscle, the nerve axons go in between all the muscle cells and stimulate all cells separately. This is for example seen in the lens of the eye.
Here, the protein-coding genes expressed in smooth muscle are described and characterized, together with examples of immunohistochemically stained tissue sections that visualize corresponding protein expression patterns of genes with elevated expression in smooth muscle.
Transcript profiling was based on a combination of two transcriptomics datasets (HPA and GTEx), corresponding to a total of 14590 samples from 54 different human normal tissue types. The final consensus normalized expression (nTPM) value for each tissue type was used for the 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)