Anterior pituitary development: Short tales from dwarf mice

Cell, Vol. 70, 527-530, August 21, 1992, Copyright 0 1992 by Cell Press

Anterior Pituitary Development: Short Tales from Dwarf Mice

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Jeffrey W. Voss’ and Michael G. Rosenfeld*t ‘Eukaryotic Regulatory Biology Program tHoward Hughes Medical Institute Department of Medicine University of California, San Diego School of Medicine La Jolla, California 92093-0648

derm and neuroectoderm in the primitive head (Schwind, 1928), implying that cell-cell contact may be an inductive event. Coincident with these cell-cell contacts on embryonic day 11 (ell) in the rat, the first known anterior pituitary marker, the aGSU transcript, is restricted to most or all cells in a single layer of epithelium in a clear posterioranterior

gradient

(Simmons

et al., 1990), suggesting

an

asymmetry of the inducing factor(s). Unanswered quesDefining the mechanisms by which specialized cells arise and generate organs containing functionally diverse cell types is of fundamental importance in understanding development. Study of the five distinct cell types of the mammalian anterior pituitary gland that exert distinct functions in homeostatic control in vertebrates has provided insight into these mechanisms.

Together,

the five cell types regu-

late positive and negative feedback control from the brain (via the hypothalamus) to peripheral endocrine organs, such as the thyroid, adrenal, and gonads (Figure 1). These cells coordinate central nervous system regulation of vital processes, including metabolism, growth, reproduction, and behavior. Rathke’s pouch, the pituitary precursor, becomes committed to organ development several days before the expression of markers of individual cell types within the mature gland. Between the time of organ commitment and organ maturation, a series of cell type-specific differentiation and proliferation events occur. More complete molecular details of these events will provide an understanding of the general principles of mammalian organogenesis,

as well as the specific

regulation

tions are whether

organ commitment

precedes

cell-ceil

contact and whether the inducing signal is extracellular or membrane bound. A parallel issue is whether there are cells that fail to express the aGSU marker at this stage that serve as precursors of certain anterior pituitary cell types. Commifmenf to Organogenesis and Cell Type Differentiation While lineage analysis is not available for pituitary cell types, organ culture experiments have suggested that the cells in the pituitary anlage are committed to organ development as early as e12 in the rat (Bggeot et al., 1982). This approach

has provided

evidence

for presentation

of

several morphogenic substances to the developing pituitary at different times, resulting in the sequential appearance of distinct cell types. The first cell type to appear in vivo and in organ culture is the corticotroph. The appearance of corticotrophs independent of the composition of the culture medium (e.g., Bggeot et al., 1982) argues that triggering of corticotroph

of pitu-

itary development. Ontogeny of Anferior Pituitary Cell Types The five cell types in the mature anterior pituitary gland are defined by the trophic factors that they synthesize and secrete.

Corticotrophs

produce

adrenocorticotrophin

(ACTH) by proteolytic processing of proopiomelanocortin (POMC), regulating adrenal cortex production of glucocorticoids;

thyrotrophs

synthesize

thyroid-stimulating

hor-

mone (TSH), which regulates thyroid gland growth and hormone hormone

production; gonadotrophs (LH) and follicle-stimulating

produce luteinizing hormone (FSH), reg-

ulating gonadal function; somatotrophs produce growth hormone (GH), regulating linear growth; and lactotrophs synthesize prolactin (Prl), which regulates milk production. TSH, LH, and FSH are heterodimeric glycoproteins consisting of a common a subunit (aGSU) and a distinct b subunit.

Because these five cell types arise in a precise temporal and spatial pattern (Figure 2), a critical question is: what triggers the appearance of each cell type from an apparently homogeneous

primordium?

The anterior

pituitary

it-

self arises from ectodermal cells adjacent to the anterior neuropore, which demarcates the limit of the developing neural tube. Involution of these ectodermal cells forms a layer of cells that makes contact with the neuroectoderm that gives rise to the hypothalamus. This contact is the only region of mesodermal

incompetence

between

ecto-

Figure 1. Selected Feedback Regulatory Loops in the Neuroendocrine System A few of the many regulatory pathways are shown schematically, including both hypophysiotrophic regulators from specialized hypothalamic neurons, which are delivered via a portal capillary system, and feedback from target organs.

EMBRYONIC DAY

“0;;;;

TARGETWiG DNA SEGUENCES

CELL-TYPE ONTOGENY

410 ?jl

ell

FACTOR ONTOGENY

y

//-? G Thyrotropes and el 1 pituitary precursors

-706 -

TEF

e14

Figure 2. Spatial, Anatomical, and Ontogenic Relationships in the Developing PituitaryHypothalamic Axis Schematic representations of the pituitary gland are shown on the days that each cell type can first be detected. Cell types are indicated on the day and in the position that they are born (although each persists throughout the life of the animal). The hormone gene DNA sequences that target cell type-specific expression in transgenic animals are indicated. Sinding sites for Pit-i and TEF are indicated in Prl, GH, PTSH, and Pit-l promoters.

e15

FSH .LH __,I

e16

birth

E2

LACTOTROPE

PROLIFERATION

differentiation takes place very early and largely independent of influences outside the organ. In contrast, appearance of gonadotrophs, somatotrophs, and lactotrophs in organ culture requires the addition of hormones, illustrating the potential role of endocrine and paracrine processes in regulating the differentiation of particular cell types. For example, addition of insulin-like growth factor 1 (IGF-l), cortisol, or thyroid hormone in various combinations results in the appearance of somatotrophs (Hemming et al., 1984). The hypothalamic regulator, gonadotropin releasing hormone (GnRH) results in the differentiation of gonadotrophs and, by mediating release of aGSU from gonadotrophs, the appearance of prolactin-expressing lactotrophs (Begeot et al., 1984). Organ culture experiments have not distinguished whether hormones activate the differentiation of cell types, increase expression of already activated, trophic factorencoding genes, or trigger the expansion of an undetectably small population of previously differentiated cells. Thus, a significant question in pituitary development remains whether the committed Rathke’s pouch tissue consists of a mixture of incompletely differentiated but totipotent cells, or if it contains small numbers of partially committed cells that await proliferative cues. Recent anal-

ysis of inheritable pituitary dwarfism described below has, in part, resolved this issue. Organ culture experiments have, however, established that delivery of agents to the developing pituitary at different times can have profound effects on the types of cells that develop. Development and Regulation of Cell-Specific Proliferation Individual cell types arise within restricted regions of the developing gland (Simmons et al., 1990; Dubois and Hemming, 1991; Figure 2). This pattern of pituitary development is conserved throughout vertebrates. In the mature animal, the five mature cell types are more homogeneously distributed throughout the anterior pituitary. This could reflect either a loss of homophilic interactions between cell types or the action of migration-inducing factors. Fish, in contrast, preserve a stratified distribution of pituitary cell types. The variegated appearance of differentiated cell types within restricted portions of the anterior pituitary may indicate that they arise from progenitor cells (or even clones) induced to differentiate or proliferate by agents from other tissues or the pituitary itself. Proliferation regulates aspects of pituitary physiology. On e14, the most rapidly replicating cells in the anterior pituitary gland are those that maintain contact with neu-

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roectoderm. Thyrotrophs and corticotrophs appear to arise in regions that exhibit limited cellular proliferation. In the neonatal pituitary, however, replicating cells appear to be distributed throughout the organ (Ikedaand Yoshimoto, 1991). In the adult, the relative numbers of each of the five cell types vary considerably depending on the endocrine state of the individual, and each cell type proliferates in response to hormones and hypothalamic trophic factors. Best characterized are the somatotrophs, whose proliferation is regulated by intracellular CAMP levels, which can be increased by the binding of growth hormone-releasing factor to its receptor (Billestrup et al., 1988). This model is supported by pituitary adenomas that express a mutated, constitutively active form of the as subunit of the heterotrimeric G, protein (Landis et al., 1989) and somatotroph hyperplasia in transgenic animals expressing chimeric cholera toxin genes under control of the growth hormone promoter (Burton et al., 1991). Moreover, expression of a dominant negative, mutant CREB gene results in the generation of dwarf transgenic animals nearly devoid of somatotrophs(Strutherset al., 1991). Incontrast, estrogen increases lactotroph proliferation as well as prolactin gene expression (Carbajo-Perez and Watanabe, 1990, and references therein), and the appearance of estrogen receptor in the anterior pituitary coincides with a rapid increase in the number of lactotrophs (Slaughbaugh et al., 1981; Figure 2). Whether the mitogenic agents that affect adult pituitary cell proliferation are identical to those that affect the initial proliferation during ontogeny remains to be addressed. Genetic Approaches to Cell Lineage Relationships Further insight into the differentiation of pituitary cell types has been provided bygeneticablation experiments. These experiments are feasible because upstream regulatory sequences have been defined that are sufficient to promote regulated, cell-specific gene expression (e.g., Drouin et al., 1989; Crenshaw et al., 1989; Windle et al., 1990). Because many cells coexpress growth hormone and prolactin (Hoeffler et al., 1985), it was suspected that somatotrophs and lactotrophs are derived from a common lineage. Evidence for this has been provided by the dwarf phenotype and virtual absence of growth hormone-expressing cells in transgenic mice expressing a growth hormone-diphtheria toxin fusion gene (Behringer et al., 1988). Most lines also contained vastly reduced lactotroph populations. Expression of a viral thymidine kinase gene fused to either the growth hormone or prolactin promoters (Borrelli et al., 1989) has permit “induced” cytotoxicity in cells replicating DNA by administration of fluoroiodoarabinouridine at various times during the development of transgenic animals. The growth hormone-thymidine kinase transgene is lethal to nearly all somatotrophs and lactotrophs. Together, these data suggest that most lactotrophs arise from cells that have, at some point, expressed the growth hormone gene. The phenotypes of these dwarf animals suggest that the other pituitary cell types arise independently of cells that express growth hormone. Experiments using aGSU promoter-diphtheria toxin fusions have demonstrated that

gonadotrophs can be ablated with a reduction only in lactotrophs and very little effect on other pituitary cell types (Kendall et al., 1991). This may reflect decreased levels of aGSU or estrogen due to the elimination of gonadotrophs. More importantly, these results imply either that all pituitary cell types do not arise from the aGSU-expressing cells in the el 1 pre-Rathke’s pouch or that cells expressing aGSU early in development use regulatory sequences distinct from those used by mature gonadotrophs, such as those required for aGSU expression in thyrotrophs. However, developmentally regulated promoter switching has not yet been described for any pituitary-specific gene. Genetic Evidence for Cell-Specific Transcription Factors as Developmental Regulators In cell culture systems, both the growth hormone and prolactin genes are activated by the pituitary-specific POU domain transcription factor Pit-l (or GHF-1) (Ingraham et al., 1990; Fox et al., 1990; Castrillo et al., 1991) which is ultimately present in somatotrophs, lactotrophs, and thyrotrophs. Analyses of transgenic animals indicate that the expression of Pit-i target genes is selectively restricted in part by the DNA sequence context of the enhancers of the target genes (Crenshaw et al., 1989). However, mouse genetics has provided direct evidence for the developmental role of the tissue-specific transcription factors. A variety of developmental mutants have been identified by their dwarf phenotypes. Among these are the Snell, Jackson, and Ames dwarf mutants, which exhibit a virtually identical phenotype-theycontain nolactotrophs, somatotrophs, or thyrotrophs (Slaughbaugh et al., 1981) and have markedly hypoplastic anterior pituitary glands. Jackson and Snell dwarfs are allelic on chromosome 18 (Li et al., 1990; Camper et al., 1991); it has been demonstrated that both dwarf phenotypes result from mutations of the Pit-l gene (Li et al., 1990). The Jackson mutation is a rearrangement, whereas the Snell phenotype is a missense mutation in the Pit-l POU homeodomain. This crucial residue in the third (or DNA recognition) helix is conserved among all homeodomain proteins. The mutation impairs the ability of Pit-l to bind to its DNA recognition elements. Consistent with Pit-l regulating the prolactin and growth hormone genes, the onset of expression of Pit-l protein on e15.5 (Simmons et al., 1990; Dolle et al., 1990) correlates closely with that of the prolactin and growth hormone genes. However, the failure of the dwarf animals to develop thyrotrophs, which appear more than a day before detectable Pit-l gene expression, suggests that Pit-l may be involved in thyrotroph survival or maintenance, as opposed to their establishment. The hypoplastic dwarf pituitary gland containing only gonadotrophs and corticotrophs reveals that Pit-l is important for proliferation (and/ or survival) of several cell types, and one report has suggested that addition of antisense oligonucleotides against Pit-l mRNA can decrease proliferation of a rat pituitary cell line (Castrillo et al., 1991). The ability of Pit-l to activate target genes and regulate proliferation or survival may parallel the actions of uric-88 in sensory neuron development in Caenorhabditis elegans (Ruvkun and Finney, 1991). However, the expression of

thyrotroph embryonic factor (TEF), a basic-leucine repeat transcription factor, and not that of Pit-l correlates spatially and temporally with the onset of TSHb expression in the rostra1 tip of the developing gland; TEF can uniquely activate TSHf3 expression in transfection analysis (Drolet et al., 1991). Interestingly, Pit-l transcripts are not expressed in Snell and Jackson dwarfs. This may, in part, reflect Pit-l gene autoregulation that perhaps provides a”memory” for maintaining the differentiated state of two or three pituitary cell types(Chen et al., 1990; McCormicket al., 1990). Because the nonallelic Ames dwarf mutation produces a phenotype indistinguishable from that caused by the Jackson and Snell mutations in the Pit-i gene, it is tempting to speculate that this gene may function epistatically to Pit-l. Conclusions Anterior pituitary gland development involves a commitment to organogenesis by precursor cells and subsequent restriction of the initially expressed markers to specific cell types. This restriction is followed by differentiation, which is coupled to the expression of secondary markers defining the mature cell types. The appearance of the five pituitary cell types suggests the actions of inducing substances and cell-cell interactions during development. These cues activate regulatory mechanisms that are progressively refined by expression of additional activating and restricting factors, such as Pit-l. These processes that give rise to distinct cell types within the mature gland are likely to be general features of mammalian organogenesis.

Hemming, F. J., Begeot, M., Dubois, M. P., and Dubois, P. M. (1984). Endocrinology 774, 2107-2113. Hoeffler, J. P., Boockfor, F. R., and Frawley, L. S. (1985). Endocrinology 777, 187-195. Ikeda, H., and Yoshimoto,

T. (1991). Cell Tissue Res. 263, 41-47.

Ingraham, H., Albert, V. R., Chen, R., Crenshaw, E. B., Elsholtz, H. P., He, X., Kapiloff, M. S., Mangalam, H. J., Swanson, L. W., Treaty, M. N., and Rosenfeld, M. G. (1990). Annu. Rev. Physiol. 52.773-791, Kendall, S. K., Saunders, T. L., Jin, L., Lloyd, R. V., Glode, L. M., Nett, T. M., Keri, R. A., Nilson, J. H., and Camper, S. A. (1991). Mol. Endocrinol. 5, 2025-2038. Landis, C. A., Master, S. B., Spada, A., Pace, A. M., Bourne. H. R., and Vallar, L. (1989). Nature 340, 692-696. Li, S., Crenshaw, E. B., Rawson, E. J., Simmons, D. M., Swanson, L. W., and Rosenfeld, M. G. (1990). Nature 347, 528-530. McCormick, A., Wu, D., Castrillo, J. L., Dana, S., Strobl, J., Thompson, E. B., and Karin, M. (1990). Nature 345, 829-833. Ruvkun. G., and Finney, M. (1991). Cell 64, 475-478. Schwind, J. L. (1928). Am. J. Anat. 47, 295-319. Simmons, D. M., Voss, J. W., Ingraham, H. A., Holloway, J. M., Broide, R. S.. Rosenfeld, M. G., and Swanson, L. W. (1990). Genes Dev. 4, 695-711. Slaughbaugh, M. B., Lieberman. M. E., Rutledge, J. J., and Gorski, J. (1981). Endocrinology 709, 1040-1046. Struthers, R. S., Vale, W. W., Arias, C., Sawchenko, miny, M. R. (1991). Nature 350, 622-824.

P. E., and Mont-

Windle, J. J., Weinger, R. I., and Mellon, P. L. (1990). Mol. Endocrinol. 4, 597-603. Note Added

In Proof

The present address of Jeffrey Voss is BASF Bioresearch Corporation, 195 Albany St., Cambridge, Massachusetts 02139.

References Begeot, M., Dubois, M. P., and Dubois, P.M. (1982). Neuroendocrinology 35, 255-264. Begeot, M., Hemming, F. J., Combarnous, Y., Dubois, M. P., Aubert. M. L., and Dubois, P. M. (1964). Science 226, 566-566. Behringer, R. R., Mathews, L. S., Palmitter, Ft. D., and Brinster, Ft. L. (1986). Genes Dev. 2,453-461. Billestrup, N., Swanson, L. W., and Vale, W. (1986). Proc. Natl. Acad. Sci. USA 83, 8654-6857. Borrelli, E., Heyman, R. A., Arias, C., Sawchenko, R. M. (1989). Nature 339, 538-541.

P. E., and Evans,

Burton, F. H., Hasel, K. W., Bloom, F. E., and Sutcliffe, J. G. (1991). Nature 350, 74-77. Camper, S. A., Saunders, T. L., Katz, R. W., and Reeves, R. H. (1991). Genomics 8, 586-590. Carbajo-Perez, 333-338.

E., and Watanabe,

Y. G. (1990). Cell Tissue Res. 267,

Castrillo, J. L., Theill, L. E., and Karin, M (1991). Science 253, 197199. Chen, R., Ingraham, H. A., Treaty, M. N., Albert, V. R., Wilson, L., and Rosenfeld, M. G. (1990). Nature 346, 583-586. Crenshaw, Rosenfeld,

FOX, S. R., Jong, M. T. C., Casanova, J., Ye, S. F., Stanley, F., and Samuels, H. H. (1990). Mol. Endocrinol. 4, 1069-1080.

E. C. C., Kalla, K., Simmons, D. M., Swanson, M. G. (1989). Genes Dev. 3, 959-972.

L. W., and

Doll& P., Castrillo, J. L., Theill, L. E., Deerinck, T., Ellisman, M., and Karin, M. (1990). Cell 60, 809-820. Drolet, D. D., Scully, K. M., Simmons, D. M., Swanson, L. W., Wegner, M.. Chu, K., and Rosenfeld, M. G. (1991). Genes Dev. 5, 1739-1753. Drouin, J., Nemer, M., Charron. J., Gagner, J. P., Jeannotte, Sun, Y. L. (1989). Genome 37, 510-519. Dubois, P. M., and Hemming, 2-20.

L., and

F. J. (1991). J. Elect, Micro. Tech. 79,