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The Cellular Biology of Erythropoietin Receptors

Queen’s University Belfast, Belfast City Hospital, Belfast, Northern Ireland

Correspondence: Terry Lappin, Ph.D., Queen’s University Belfast, Belfast City Hospital, U Floor, Tower Block, Lisburn Road, Belfast BT9 7AB, Northern Ireland. Telephone: 44-0-2890-329241, ext 2013; Fax: 44-0-2890-263927; e-mail: t.lappin{at}qub.ac.uk

	LEARNING OBJECTIVES

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 
After completing this course, the reader will be able to:

1. Explain the structure and functions of erythropoietin.
   
2. Distinguish the multiple activities of erythropoietin in the human body.
   
3. Recognize the potential role of erythropoietin receptors in targeting tumors.
   

	ABSTRACT

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 
Long thought to act only as a hormone that was the primary regulator of red blood cell production, erythropoietin is now known to have a whole spectrum of activity. Organs such as the brain, ovary, oviduct, uterus, and testis have erythropoietin receptors. Because erythropoietin receptors exist in the paracrine and autocrine systems, as well as the hormonal system, the beneficial effects of administering human erythropoietin are likely to extend beyond its effect in raising hematocrit. It may even be possible in the future to use the receptors to target a drug to a tumor without damaging the surrounding tissue.

Key Words. Erythropoietin receptors • Recombinant erythropoietin • Neoplasms

	INTRODUCTION

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 
Adult humans produce approximately 2.3 million red blood cells every second, or 138 million every minute. The main regulator of that process is erythropoietin, a glycoprotein hormone that circulates at about one hundredth of the concentration of most other hormones in the body [1, 2]. Erythropoietin is produced in the kidneys. It circulates in the plasma and induces red cell production in the bone marrow [3], where it binds to erythroid progenitor cells. Cell culture studies have identified two classes of erythroid progenitor cells, BFU-E and colony forming units-erythroid (CFU-E) (Fig. 1). Both types of cell have receptors for erythropoietin on their surfaces. When erythropoietin binds to its receptors on BFU-E cells, they proliferate into CFU-E (proerythroblasts). Proerythroblasts are exquisitely sensitive to erythropoietin. They proliferate and develop into erythroblasts and then reticulocytes that enter the peripheral circulation where they mature into red blood cells.

View larger version (46K): [in this window] [in a new window]   Figure 1. The endocrine action of erythropoietin (EPO) depends on a feedback loop, as proposed by Erslev and Gabuzda [3]. The author is grateful to Professor A. Peter Maxwell for the illustration.

Investigators are now beginning to understand what actually happens at the cellular level when erythropoietin binds to its receptor, and it is much more complicated than originally thought. For example, there are numerous built-in mechanisms to ensure that the stimulatory effect of erythropoietin on erythroid progenitor cells is appropriate. Shortly after erythropoietin binds to its receptor and initiates the signaling cascade that leads to cell proliferation and prevention of apoptosis, its effect is attenuated by intracellular molecules such as suppressors of cytokine signaling, which actually prevent uncontrolled proliferation of red blood cells. Basically, erythropoietin docks on the outside of the cell and sends messages to the nucleus, which result in the proliferation of the cells. More cells develop, differentiate, and are prevented from experiencing natural cell death (apoptosis) by the presence of erythropoietin. Thus, all the major processes involved in red blood cell development, survival, growth, and maturation are maintained by erythropoietin binding to the receptors of the maturing erythroid cells.

	STRUCTURE OF ERYTHROPOIETIN

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 
Erythropoietin is a glycoprotein molecule composed of 165 amino acids and four carbohydrate groups. The primary structure is shown in Figure 2. An important structural feature of erythropoietin is that it has two disulphide bonds, one linking the cysteine at amino acid 6 with the cysteine at amino acid 161, and the other linking cysteines 29 and 33. The former is functionally more important, because it acts as a tether, ensuring that the whole molecule is held in the correct shape for binding to the erythropoietin receptor. If this bond breaks, the molecule loses its biological activity. From the structure of the erythropoietin molecule, and from experimental evidence, one can infer that molecules of erythropoietin aggregate together through a process known as hydrophobic interaction. When this happens to any degree, perhaps as a result of improper storage, erythropoietin becomes significantly less potent.

View larger version (47K): [in this window] [in a new window]   Figure 2. The primary structure of erythropoietin showing the circulating form of 165 amino acids. Two disulfide bonds tether the molecule together between cysteines 29 and 33 and cysteines 6 and 161. Three N-linked sugars are present at asparagines 24, 38, and 83, and one O-linked sugar is present at serine 126.

	ERYTHROPOIETIN RECEPTORS

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 
The erythropoietin receptor actually consists of an extracellular, a transmembrane, and an intracellular domain. A single erythropoietin molecule binds to two receptors on the cell surface (Fig. 3). As a consequence, tyrosines located in the intracellular domain are phosphorylated, initiating an intracellular signaling cascade that regulates gene expression in the nucleus, which in turn controls cell survival, proliferation, and differentiation.

View larger version (36K): [in this window] [in a new window]   Figure 3. Erythropoietin (EPO) binds to two receptors. The author is grateful to Professor A. Peter Maxwell for the illustration.

Immunohistochemistry, a technique that uses antibodies to detect specific proteins, has been used to identify erythropoietin receptors on cell membranes in various tissues. Another technique, in situ hybridization, can be used to detect erythropoietin receptor mRNA in individual cells. These techniques can be used in tandem to confirm the presence of both erythropoietin receptor mRNA and protein in the same cell. This approach rules out the possibility that a nonspecific interaction has occurred between an anti-erythropoietin receptor antibody and a cell membrane protein that resembles the erythropoietin receptor.

	ACTION OF ERYTHROPOIETIN

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 
The endocrine action of erythropoietin originates in the kidney. This erythropoietin acts as a hormone; it is produced in one tissue and is transported in the plasma to a target tissue. In the brain, however, there are cells that make erythropoietin that binds to erythropoietin receptors on nearby cells; this is called a paracrine system. In addition, some of the cells in the brain make their own erythropoietin. This is an example of an autocrine process. Thus, it appears that there are three really distinct systems: A) the endocrine, or hormonal, system; B) the paracrine system, which functions locally; and C) the autocrine system, in which the cell that is going to use the erythropoietin actually makes it for itself. The relative importance of these three systems remains to be determined.

The size of the erythropoietin molecule produced in brain tissue is smaller than erythropoietin made in the kidney, because some of the sialic acid groups are missing from the carbohydrate chains [4]. In the paracrine system in the brain, erythropoietin binds to receptors on adjacent cells. It, therefore, does not enter the circulation and is not susceptible to removal by the liver, rendering the addition of sialic acid groups unnecessary.

	ERYTHROPOIETIN IN THE FUTURE—TREATING MORE THAN FATIGUE?

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 
Repeating some work that had been done recently by Acs et al. [5], we looked at erythropoietin receptors in breast cancer tissue in our own laboratory [P. Maxwell, unpublished data]. Using immunohistochemistry, we found that erythropoietin receptors were present in tumor cells but were absent from the surrounding normal breast tissue. This is significant, because it may be possible to use a tumor’s erythropoietin receptors to target a drug to a tumor and not damage the surrounding tissue. This concept appears to have potential for future exploitation.

Another potential therapeutic intervention for patients with cancer involves the intracellular signaling cascade. Its mechanism of action in the erythroid cells has been extensively investigated. If future investigations can determine the signaling cascade in cancer cells, it may be possible to direct therapy at a specific target in the cancer cells while sparing the normal cells.

A major question for the future is: Does erythropoietin have an antitumor effect, apart from raising the hematocrit and increasing the oxygen available? Also, in view of the discovery of erythropoietin receptors in tumor cells, another important question arises: Does the erythropoietin receptor or its signaling pathway in tumor cells represent a real chemotherapeutic target? Clearly, further work is required to address these important issues in cancer chemotherapy.

	REFERENCES

Top Learning Objectives Abstract Introduction Structure of Erythropoietin Erythropoietin Receptors Action of Erythropoietin Erythropoietin in the... References

 

1. Lappin TR, Maxwell AP, Johnston PG. EPO’s alter ego: erythropoietin has multiple actions. STEM CELLS 2002;20:485–492.[Abstract/Free Full Text]
2. Maxwell AP. Novel erythropoiesis-stimulating protein in the management of the anemia of chronic renal failure. Kidney Int 2002;62:720–729.[CrossRef][Medline]
3. Erslev AJ, Gabuzda TG. Pathophysiology of Blood, third edition. Philadelphia: WB Saunders, 1985:28–134.
4. Masuda S, Okano M, Yamagishi K et al. A novel site of erythropoietin production. Oxygen-dependent production in cultured rat astrocytes. J Biol Chem 1994;269:19488–19493.[Abstract/Free Full Text]
5. Acs G, Acs P, Beckwith SM et al. Erythropoietin and erythropoietin receptor expression in human cancer. Cancer Res 2001;61:3561–3565.[Abstract/Free Full Text]

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This Article Abstract Full Text (PDF) CME: Take the course for this article: The Cellular Biology of Erythropoietin Receptors eLetters: Submit a response to this article Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services E-mail this article link to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints/Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Lappin, T. Search for Related Content PubMed PubMed Citation Articles by Lappin, T.