The bone marrow-specific proteome

The bone marrow transcriptome

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

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

As shown in Figure 1, 534 genes show some level of elevated expression in the bone marrow compared to other tissues. The three categories of genes with elevated expression in bone marrow compared to other organs are shown in Table 1. In Table 2, the 12 genes with the highest enrichment in bone marrow are defined.

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

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

Protein expression of genes elevated in bone marrow

In-depth analysis of the elevated genes in bone marrow using antibody-based protein profiling allowed us to visualize the expression patterns of these proteins.

The list of elevated genes (n=534) is well in line with the function of the bone marrow, as it includes an overrepresentation of proteins associated with immune response, leukocyte migration and respiration. Among the genes with the highest level of enriched expression in bone marrow (Table 2), five encode proteins with known functions in neutrophils and monocytes (AZU1, CTSG, DEFA4, ELANE and MPO). Two other elevated proteins are encoded in lymphocytes (DEFA1, DEFA1B) and four genes encode proteins for erythrocyte function (HBB, HBD; both hemoglobin proteins, GYPA, an intrinsic membrane protein and RHAG, thought to be part of a membrane channel which transports ammonium and carbon dioxide). Both neutrophils and erythrocytes reach maturity in bone marrow and are released into the bloodstream as effector cells, equipped with necessary proteins for their specialized functions. Consequently, a high level of transcription of corresponding genes takes place in bone marrow, explaining their high TS-score seen in Table 2.

Proteins with enriched expression in granulocytes

Besides erythropoietic cells and thrombocytes, polymorphonuclear leukocytes cells, and in particular cells of the neutrophil lineage, make up the majority of hematopoietic cells in bone marrow. CTSG (Cathepsin G) and DEFA4 (Defensin alpha 4) are two of the elevated genes within bone marrow. They are known to be expressed in neutrophils and involved in the defense against bacteria. PRTN3 (Proteinase-3), a neutrophil serine protease (NSPs) differs from the other two NSPs (CTSG and ELANE) in terms of function. An example being that proteinase-3 also acts as a feedback regulator in myeloid differentiation. Protein profiles for CTSG, PRTN3 and DEFA4 show strong staining of granulocytes.

CTSG
PRTN3
DEFA4

Proteins with enriched expression in monocytes

The granulocytic mast cells and the agranulocytic macrophages are present in lower numbers than neutrophils in bone marrow. MCEMP1, a fairly uncharacterized gene found to encode a single-pass transmembrane protein expressed in human mast cell, displays a group enriched expression in bone marrow, along with lung and appendix. The RNA-seq data is supported by immunohistochemistry, with positivity in subsets of cells in bone marrow and appendix, as well as in alveolar macrophages in the lung.

MCEMP1 - bone marrow
MCEMP1 - lung
MCEMP1 - appendix

Gene expression shared between bone marrow and other tissues

There are 135 group enriched genes expressed in bone marrow. 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 bone marrow, compared to all other tissues.

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

Among the group enriched genes in bone marrow we found two genes, S100A12 (S100 calcium binding protein A12) and IGLL1 (Immunoglobulin lambda like polypeptide 1) display an enriched expression in lymphoid tissue and testis, respectively. S100A12 is a fairly well characterized calcium-binding proinflammatory protein constitutively expressed in both neutrophils and macrophages, and secreted by activated neutrophils. Immunohistochemical staining of S100A12 is in concordance with the RNA-seq data and literature, showing a strong positivity in bone marrow and peripheral lymphoid tissues.

S100A12 - bone marrow
S100A12 - spleen

IGLL1 is a less known gene that seems to encode a protein expressed in pre-B-cells and pro-B-cells. Immunohistochemistry displays cytoplasmic staining in bone marrow and testis, which is supported by RNA-seq data.

IGLL1 - bone marrow
IGLL1 - testis

Bone marrow function

The main functions of the bone marrow are to maintain constant levels of the different blood cell types in the peripheral blood, i.e. producing erythrocytes, leukocytes and thrombocytes. Bone marrow also contributes to the degradation of aged erythrocytes, along with the liver and spleen.

Bone marrow histology

Bone marrow is divided into red and yellow regions, caused by a predominance of either hematopoietic-rich (red) or adipose-rich (yellow) tissue. Red marrow consists of a highly vascularized stromal network containing pluripotent and committed stem cells of all hematopoietic lineages, i.e. erythrocytes, leukocytes, thrombocytes. While erythrocytes and leukocytes develop from stages of precursors, thrombocytes, small blood cell fragments involved in clotting, originate from giant marrow cells called megakaryocytes. In contrast, yellow marrow contains mesenchymal stem cells which differentiate into several stromal lineages, such as chondrocytes, osteoblasts, fibroblasts and adipocytes.

At birth and until around the age of seven, all human marrow is red, as the need for new blood formation is high. Adipose tissue gradually replaces the red marrow, which in adults is mainly found in flat bones, such as the vertebrae, ilium, sternum and cranium as well as at the epiphyseal ends of the long bones of the arm and leg.

Figure 3. Schematic view of bone marrow tissue. Attribution: By Mysid [Public domain], via Wikimedia Commons. Source

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

Background

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

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

Andersson S et al., The transcriptomic and proteomic landscapes of bone marrow and secondary lymphoid tissues. PLoS One. (2014)
PubMed: 25541736 DOI: 10.1371/journal.pone.0115911

Histology dictionary - bone marrow