The esophagus-specific proteome

The esophagus transcriptome

Transcriptome analysis of the esophagus 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 esophagus compared to other tissues. Elevated expression includes three subcategory types of elevated expression:

• Tissue enriched: At least four-fold higher mRNA level in esophagus 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 esophagus 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 esophagus compared to other tissues. As evident in Table 1, all genes elevated in esophagus 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 esophagus as well as in all other tissues. (B) The distribution of all genes across the six categories, based on transcript detection (NX≥1) in esophagus as well as in all other tissues.

As shown in Figure 1, 311 genes show some level of elevated expression in the esophagus compared to other tissues. The three categories of genes with elevated expression in esophagus compared to other organs are shown in Table 1. In Table 2, the 5 enriched genes are defined.

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

Table 2. The 5 genes with enriched expression in esophagus. "Tissue distribution" describes the transcript detection (NX≥1) in esophagus as well as in all other tissues. "mRNA (tissue)" shows the transcript level in esophagus as NX values. "Tissue specificity score (TS)" corresponds to the fold-change between the expression level in esophagus and the tissue with second highest expression level.

Protein expression of genes elevated in esophagus

In-depth analysis of the elevated genes in esophagus using antibody-based proteomics allowed us to create an overview of the localization of the corresponding proteins. A large number of these proteins have functions related to squamous differentiation and are thus often also shared with other tissue types that are composed of squamous epithelia.

Proteins specifically expressed in esophageal epithelia

The inner lining of the esophagus is made up by glycoprotein-rich mucosal squamous epithelium that lacks an outer layer of cornified cells (as in the skin). Like most squamous epithelia, the esophagus express a variety of keratin intermediate filament proteins whose function is to provide structural integrity between the cells. Among structural proteins, Keratin 4 (KRT4), -6 (KRT6A, KRT6B and KRT6C) -13 (KRT13), and -32 (KRT32) showed high enrichment together with the calcium-binding proteins cornulin (CRNN) and S100A14. KRT13 is primarily expressed in the mucosal epithelia, as MUC21, which is observed in esophageal epithelial cells. An interesting enriched protein is the alcohol-degrading enzyme ADH7, which is observed in mucinous epithelial cells of the esophagus and stomach.

KRT6A
CRNN
ADH7

Proteins specifically expressed in esophageal muscle

Among the genes that show enrichment in the esophagus, but do not show protein expression in the epithelial cells, are two muscle-specific genes: MYH3, MYH8, and the transcription factor NKX6-1. Whereas both MYH3 and MYH8 are well-known muscle-specific genes that are group enriched in esophagus and skeletal muscle, NKX6-1 appears to be specifically expressed in muscles in the esophagus and is previously not described in this tissue.

MYH3
NKX6-1

Gene expression shared between esophagus and other tissues

There are 57 group enriched genes expressed in esophagus. 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 esophagus, compared to all other tissues.

In order to illustrate the relation of esophagus 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 esophagus enriched and group enriched genes connected to their respective enriched tissues (grey circles). Red nodes represent the number of esophagus 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.

The esophagus shares a great amount of expressed genes (n=34) with tongue, which is a tissue with highly similar squamous epithelial structure as the esophagus. Many of these tongue/esophagus group enriched genes belong to gene families known to be important for normal squamous epithelial function as for example transglutaminases; transglutaminase1 (TGM1), -3 (TGM3) and keratins; keratin4 (KRT4), -6A (KRT6A), -6B (KRT6B) and -78 (KRT78).

TGM3 - esophagus
KRT4 - esophagus
KRT78 - esophagus

Some lymfoid tissues, like tonsil, have squamous epithelium components (an addition to its lymphocyte containing centers) and a lot of expressed genes related to squamous epithelium are shared between esophagus and lymphoid tissues (n=32). Examples of the esophagus and lymfoid tissue group enriched proteins include calcium dependent glycoprotein Desmocollin 3 (DSC3) and BARX2, a transcription factor related to genes controlling cell adhesion.

DSC3 - esophagus
BARX2 - esophagus

DSC3 - tonsil
BARX2 - tonsil

Esophagus function

The esophagus is the gastrointestinal canal that connects the mouth with the stomach. In contrast to the rest of the digestive system, the esophagus does not have any absorptive or digestive functions. Anatomically, it is continuous with the back of the oral cavity and pharynx and runs downward through the diaphragm for approximately 20-30 cm until it reaches the stomach.

When swallowing, food is pressed from the mouth and pharynx into the esophagus. The swallowing reflex then opens the upper esophageal sphincter muscle to allow entry of food to the esophagus, and the epiglottis folds down to prevent food from entering into the trachea and respiratory organs. The smooth muscles lining the length of the esophagus then contract rhythmically to help push the food towards the lower esophageal sphincter muscle that opens to allow entry of food to the stomach. Both the upper and lower sphincter muscles are constricted by default unless swallowing/vomiting. The lower sphincter muscles also protect the esophagus from the acidic contents and digestive enzymes of the stomach.

Esophagus histology

The esophagus has the same general gross anatomical and histological organization as the rest of the gastrointestinal tract with an outer muscular layer, a submucosa, a muscularis mucosa, followed by a lamina propria which in the case of the esophagus consists of a stratified squamous mucinous epithelium. However, since the esophagus is located outside of the abdominal cavity it has no mesothelial covering. Instead, the outermost layer is covered by connective tissue, so called adventitia.

The innermost part is the esophageal epithelium, which has a quite rapid turnover of cells due to the continuous wear and tear of food ingestion. Like most epithelial tissues, cell renewal takes place in the basal part of the epithelium, and as new cells are generated older cells lose contact with the basal membrane and are pushed towards the surface. At the beginning, cells close to the basal layer appear columnar with round nuclei. As these cells mature and detach, they are pushed towards the apical layer of the epithelium and gradually differentiate into flattened and tightly coupled cells.

From the inside and out, the squamous epithelium rests on the lamina propria that consists of loose connective tissue and focal lymphocytes. After this layer comes the lamina muscularis mucosae composed of smooth muscle cells, followed by the submucosal layer, which is composed of loose connective tissue containing mucus secreting glands, small blood vessels and lymphocytes. After the submucosa comes the tunica muscularis that is composed of an inner layer of circular muscles, followed by externally located longitudinal muscle fibers. In the third of the esophagus that is closed to the mouth, the external layer is composed of skeletal muscle, the middle third contains a mixture of smooth and skeletal muscle, and the third closest to the stomach contains only smooth muscle.

The histology of human esophagus including detailed images and information can be viewed in the Protein Atlas Histology Dictionary.

Background

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

Transcript profiling was based on a combination of three transcriptomics datasets (HPA, GTEx and FANTOM5, corresponding to a total of 483 samples from 37 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, 2015. Tissue-based map of the human proteome. Science
PubMed: 25613900 DOI: 10.1126/science.1260419

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

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

Histology dictionary - the esophagus