The settlement of melanoblast and the generation of pigment

After dorsolaterally migration, the melanoblast finds its destinations; and settled down surrounded by epithelial cells. What attracts melanoblast to settle in these particular regions in molecular level?

The role of the apical ectodermal ridge and of fibroblast growth factors FGF-2 and FGF-4 and of the insulin-like growth factor I (IGF-I) in the control of the migration of epidermal melanoblast into wing buds was investigated using quail–chicken chimeras. As the results, both the apical ectodermal ridge and the growth factors invariably caused the migration of epidermal melanoblasts towards them. Furthermore, FGF-2 and IGF-I and to a lesser extent FGF-4 play a decisive role in directing the migration of epidermal melanoblasts within chicken wing buds and are likely to be involved in the molecular cascade by means of which the apical ectodermal ridge controls the migration of epidermal melanoblasts. Although IGF-1 acts as upstream regulator of FGFs; FGFs have been shown to promote survival and proliferation of melanoblasts, further study has shown that the proliferation of melanoblasts can also be promoted by IGF-I, independent with FGFs (Schofer, et al., 2001).

Reaching to the destination is not sufficient for melanoblast settlement. As a kind of steam cell, melanoblast needs a population of cells (known as a “niche”) surrounding it to keep its capacity to self-renew and generate differentiated progeny (Nishimura, et al., 2002). At specific cutaneous sites, epithelial cells use Foxn1 (a transcription factor) to recruit melanocytes and induce their own pigmentation. Foxn1 thus defines a distinct cell population that ultimately controls the targeting of pigment in the skin (Weiner, et al., 2007).

After settlement, the melanoblast will start to exert its role, differentiated into melanocyte and generate pigmentation.

Mammals develop most of their coloration through a system comprised of two types of cells (reviewed by Slominski et al., 2004), referred to here as pigment donors and recipients. The pigment donors are melanocytes, which synthesize melanin in distinct organelles called melanosomes. The pigment recipients are epithelial cells, which acquire and hold most cutaneous melanin. As the system forms, each melanocyte extends dendrites and contacts multiple epithelial cells, creating a ‘‘pigmentary unit.’’ Melanosomes are then transported along the dendrites and into the epithelial cells, which may internalize the melanosomes via phagocytosis (Weiner, et al., 2007).

But the relationship between the donor melanocytes and the recipient epithelial cells are not changeless. Some differentiated melanocytes can return to the “niche” and become stem-cell again (Nishimura, et al., 2002).

Studies of pigment generation in melanocytes always focus on the function of the three transcription factors MITF, PAX3, and SOX10. The pathways of these genes interact to regulate crucial aspects of melanocyte development and function. Microphthalmia-associated Transcription Factor (MITF) has been termed the “melanocyte master regulator”, because it plays such a central role in melanocyte development and function. It is required for melanocyte differentiation and survival, activates transcription of melanogenic enzymes and melanogenic proteins, and is associated with melanoma progression. MITF also governs numerous other cellular functions in the melanocyte, including environmental response, cell survival, cell motility, and cell cycle progression. MITF itself undergoes complex post-transcriptional regulation, including phosphorylation, sumoylation, ubiquitination, and caspase cleavage. Paired box gene 3 (PAX3) has a broader expression pattern than MITF, regulating neural tube closure, early development of myoblast and neural crest lineages, and the formation of nervous, muscular, cardiovascular and melanocyte systems. PAX3 regulation of early neural crest development appears to maintain neural crest stem cell properties via inhibition of apoptosis, and PAX3 has been proposed to inhibit apoptosis in melanoma. In melanocytes, PAX3 activates transcription of MITF and plays a crucial role in maintaining melanocyte stem cells. SRY-box containing gene 10 (SOX10) regulates specification of neural crest-derived melanocytes, neurons, and glia. SOX10 strongly activates MITF and regulates expression of melanogenic enzymes. Upstream regulation of SOX10, as well as additional downstream targets, interacting factors, and posttranslational modifications are only beginning to be ascertained. In summary, MITF, PAX3, and SOX10 serve to illustrate that the extensive data currently known on cellular processes governing melanocyte biology will provide an ideal foundation for future systems biology analyses in these cells (reviewed by Baxter et al., 2009).

Citation:

Schofer, C., Frei, K., Weipoltshammer, K., and Wachtler, F. The apical ectodermal ridge, fibroblast growth factors (FGF-2 and FGF- 4) and insulin-like growth factor I (IGF-I) control the migration of epidermal melanoblasts in chicken wing buds. Anat. Embryol. (Berl.)  2001. 203, 137–146.

Emi K. Nishimura, Siobhan A. Jordan, Hideo Oshima, Hisahiro Yoshida, Masatake Osawa, Mariko Moriyama, Ian J. Jackson§, Yann Barrandonk, Yoshiki Miyachi, Shin-Ichi Nishikawa. Dominant role of the niche in melanocyte stem-cell fate determination. Nature, 2002, Vol 416, 25 April. 854-860.

Lorin Weiner, Rong Han, Bianca M. Scicchitano, Jian Li, Kiyotaka Hasegawa, Maddalena Grossi, David Lee, Janice L. Brissette. Dedicated Epithelial Recipient Cells Determine Pigmentation Patterns. Cell. 2007. 07. 024.

Slominski, A., Tobin, D.J., Shibahara, S., and Wortsman, J. Melanin pigmentation in mammalian skin and its hormonal regulation. Physiol. Rev. 2004. 84, 1155–1228.

Laura L Baxter, Stacie K Loftus, William J Pavan. Networks and pathways in pigmentation, health, and disease. Wiley Interdiscip Rev Syst Biol Med. 2009 November 1; 1(3): 359–371.

As we learnt from the text book, the four-winged Drosophila is the phenotype (a homeotic transformation of the halteres of the third thoracic segment changed into the wings normally associated with the second thoracic segement) that results from a Ubx gene mutation. But, of course, there are many insects, such as dragonflies, butterflies, and bees, which normally have four wings. Is two-winged flies evolved from four-winged ancestors, or opposite?

To investigate this question, I look up the text book of evolution. In fact, all the phylogenetic relationship of various insect groups and evidence from the fossil record indicate that the ancestor of all these winged insects was one with four wings. During the lineage leading to files, the wings of the third thoracic segment were modified into halteres. This is not surprising, at least for me. In my point of view, flies are very successful animals and should be more evolutionary than most insects. And also I think evolution of wings from halteres is harder than degeneration of halteres from wings. But, how this evolutionary event happened in history? Scientists have at least three hypotheses to explain the transition from four-winged to two-winged insects.

About 60 years ago, the first hypothesis is that the four-winged common ancestor of flies and dragonflies lacked the Ubx gene. Somewhere in the evolutionary lineage leading to flies, Ubx appeared and resulted in the transformation of the third thoracic wings into halteres. But after the development of molecular biology, it is discovered that Ubx gene is present throughout the insects, this hypothesis could be rejected.

Then, it is common to think that the Ubx gene is in all insects but expressed different between two-winged and four-winged insects. So the second hypothesis was that the evolution of two-winged from four-winged insects is caused by the mutations that affected the regulation of Ubx. But when the Ubx expression pattern was examined in a number of different insects, it was found to be highly conserved, even in wingless insects. So, the second hypothesis could also be rejected.

The third hypothesis, currently favored, focus on the downstream targets of Ubx gene. Evidence shown that in flies, various genes involved in processes such as vein formation, sensory hair growth, and cell proliferation came under the control of Ubx. Considering that the wings have veins and sensory hairs, whereas the halteres do not; the wing is far larger and contains a greater number of cells than does a haltere, it is very likely the expression of Ubx in flies’ third thoracic segment cause the formation of halteres instead of wings. But these same genes in butterflies are not regulated by Ubx, at least not in the same way.

Combining these three hypotheses and combining the basic rule of Darwinian Theory, these three kinds of mutations must be happened in history (and even nowadays), but most of them caused lethal larva, like the null mutation of Ubx gene. Some of these mutations lower the fitness of insects that might have less offspring than others, like wings are not developed in flies’ third thoracic segment nor are halteres, which make flies have trouble on flying. After one or more generation, these mutates are completely die out and we cannot see such population in wild nowadays. What we can see today is the only few of the collection of such mutations, which is rare events, but the result is the dominance of such mutates. This is the most attractive parts of evolution for me.

Citations:

Nicholas H. Barton, Derek E.G. Briggs, Jonathan A. Eisen, David B. Goldstein, Nipam H. Patel. 2007. Evolution. Cold Spring Harbor Laboratory Press

The sentence “…a small number of fetal red blood cells are seen in the maternal blood circulation.” in the text book P309 triggered my thinking weather that is one of the reasons vivipara do not have nucleated red blood cell in their circulatory system. If large amount of nucleated blood cell in the mother’s blood occasionally contact with fetus’ blood, strong immune response will kill the fetus, so under selection, only individuals with enucleated erythrocytes can survive, I guess.

In order to test my hypothesis, I started searching for information. Firstly, I found camel and giraffe’ erythrocytes have nuclear by Google. I thought if there is evidence showing their placenta structures are different from other mammals to have less possibility of maternal and fetal blood mixing, which should be a support of my hypothesis. But later, after searched for some academic articles, I found it is wrong. “All are unique among mammals, because camel’s erythrocytes are biscuit- or wafer-shaped ovals, with very little volume, and having little packed volume in whole blood (only 27-28 %). The hypothesis that they throw back to the ancestral reptiles and other lower vertebrates seemed superficially obvious, and perhaps led to the error often published that camel erythrocytes are nucleated. In fact, they are anucleate, but are unusual and novel in vertebrates.” (Long, 2007)

Furthermore, I found a credible source meaning that enucleated erythrocytes are dominant in blood in all mammal individuals including all marsupials, placental mammals, Prototheria and monotremes. Meanwhile all known non-mammalian vertebrates, including fish, amphibian, reptiles and birds, have nucleated erythrocytes (Baskurta and Meiselman, 2010). But in certain conditions, some animals will change the amount of nucleated erythrocytes or enucleated erythrocytes in their blood. Some marsupials have some circulating nucleated red blood cells as adults (Stonehouse, 1977). “Occasionally, reptilian RBC lacking nuclei are observed. These are called ‘erythroplastids,’ and their extruded nuclei found in the plasma are referred to as ‘hematogones’” (Mauro, 1997). In fetuses with intrauterine growth restriction, higher nucleated red blood cell counts at birth and longer persistence of nucleated red blood cell count are observed. Evidence shows that metabolic acidemia is in charge of causing that (Baschat, et al., 1999). These facts seem to be refutes to my hypothesis. So I decided to focus on the differences of erythropoiesis between mammal and non-mammals.

Then, I first realized, in early stage of human embryos, blood cells are not only generated by marrow. “In developing embryos, blood formation occurs in aggregates of blood cells in the yolk sac, called blood islands. As development progresses, blood formation occurs in the spleen, liver and lymph nodes. When bone marrow develops, it eventually assumes the task of forming most of the blood cells for the entire organism. ” (From Wikipedia)

What’s more, the hepatic erythropoiesis will occur in adults when excessive bleeding happened and a lot of blood cell is needed in emergency. What interested me is hepatic erythropoiesis will generate nucleated red blood cell (Baschat, et al., 1999), different from marrow. Will there be some similarities between non-mammals nucleated red blood cell? The answer seems to be no. Some net users said birds’ bones are empty inside, so they do not have marrow. They generate their blood cell by division of mature blood cells as the erythrocytes are nucleated. These net users must misunderstand something that some of the birds’ bones have marrow, and the red blood cells are produced in the bone marrow. This picture shows which bones of pigeon are empty inside and which are filled with marrow.

 

(Ornithology, Avian Circulatory System)

Finally, I found one explanation about why mammals’ erythrocytes are enucleated but non-mammalian vertebrates have nucleated erythrocytes in the aspect of evolution. But this explanation do not convince me: “Mammals, which had developed an aerobic metabolism, emerged in the Triassic, when the oxygen content in the atmosphere was by approximately 50% lower than the current level and even lower than in the Jurassic period. A drastic decrease in the total content and percentage of oxygen in the Triassic was connected with the prevalence of arid conditions on the continents. Under these conditions, mammals got rid of the nuclei in erythrocytes (having obtained enucleate and biconcave cells, where the surface area of the contained hemoglobin was larger), which led to thinner capillaries, while the biconcave shape provided a larger exchange area. Birds, which originated from more advanced reptiles, had established powerful respiratory and circulatory systems and, since they emerged at the time when the oxygen content in the Earth atmosphere approached the present level, had no need to eliminate the nuclei from their erythrocytes.” (Gavrilov, 2013)

I am wondering, in the Triassic, new coming mammals are suffering low oxygen content, why reptiles at that time are not? Why the ancestors of birds do not get rid of the nuclei in erythrocytes in Triassic? In other words, most of us have no doubt that enucleated erythrocytes are advanced than nucleated erythrocytes, but why this advanced feature only limited in mammals? I believe there should be some connection between other features of mammals and their enucleated erythrocytes. Viviparity seems not be the connection, as the evidences shown in the former part of this blog. But I will keep trying to find that. One point I am thinking is that there are two main stages of enucleated erythrocytes development in mammals; the first stage is called primitive erythropoiesis (Palis, 2014), which looks similar with the non-mammal’s erythropoiesis. Investigating their similarities and differences in molecular level, may give us some clue that how enucleated erythrocytes is evolved from nucleated erythrocytes.

Citations:

Charles A. Long. Evolution of Function and Form in Camelid Erythrocytes. Conference on Cellular & Molecular Biology – Biophysics & Bioengineering, Athens, Greece, August 26-28 (2007) 18–24.

O.K. Baskurta, H.J. Meiselmanb. Lessons from comparative hemorheology studies. Clinical Hemorheology and Microcirculation 45 (2010) 101–108.

Nicholas A. Mauro, Russel E. Isaacks. Examination of Reptilian Erythrocytes as Models of the Progenitor of Mammalian Red Blood Cells. Comp. Biochem. Physiol. Vol. 116A, No. 4, pp (1997) 323–327.

Ahmet A. Baschat, Ulrich Gembruch, Irwin Reiss, Ludwig Gortner, Chris R. Harman, and Carl P. Weiner. Examination of Reptilian Erythrocytes as Models of the Progenitor of Mammalian Red Blood Cells. Am J Obstet Gynecol. Volume 181, Number 1 (1999) 190-195.

http://en.wikipedia.org/wiki/File:Hematopoesis_EN.svg

http://people.eku.edu/ritchisong/birdcirculatory.html

Valery M. Gavrilov. Origin and development of homoiothermy: A case study of avian energetics. Am J Obstet Gynecol. Advances in Bioscience and Biotechnology, 2013, 4, 1-17.

James Palis. Primitive and definitive erythropoiesis in mammals. Am J Obstet Gynecol. FrontiersinPhysiology. January (2014) Volume5, Article3. 1-9.