Lin12 Y Let23

Inductive signal (LIN-3)

Lateral signal

▲ FIGURE 15-13 Gradient and relay signaling in C. elegans vulva development. The anchor cell (ac) sends a signal to the vulval precursor cells (P3.p-P8.p), all of which have equivalent potential to form any of three cell types—1°, 2°, 3°. The EGF-related signal, LIN-3, is received by the LET-23 receptor. Cells receiving the highest amount of signal form 1 °, cells receiving a moderate amount form 2°, and cells receiving little or none form 3°. The three cell fates are distinguished by following their subsequent patterns of cell division. The effects of the graded LIN-3 signal are further controlled by a signal relay. After the anchor cell sends LIN-3 to the nearest cell, normally P6.p, the P6.p cell then sends a different signal to its neighbors, which turns out to be a ligand for LIN-12. The demonstration of the relay effect came from genetically removing LET-23 receptor from just the P5.p and P7p cells, which prevents them from responding to the LIN-3 signal. If direct LIN-3 graded signal accounted for all the cell types, P5.p and P7p should take on the 3° fate. However, they still manage to form 2° cells; so another system must operate. This other system is the signal for LIN-12 from the P6.p cell. [Adapted from J. S. Simske and S. K. Kim, 1995, Nature 375:142.]

mally P6.p) and the 2° fate in P5.p and P7.p, which are located slightly farther away from the anchor cell (Figure 15-13a). If determination of the 2° fate depended solely on a graded LIN-3 signal, then mutant P5.p and P7.p cells lacking the receptor for LIN-3 would be expected to assume the nonvulval 3° fate. Surprisingly, when this experiment was done, the mutant cells took on their normal 2° fate (Figure 15-13b). The most likely explanation of these results is that, when the P6.p cell takes on the 1° fate, it sends out a different signal that normally works with moderate levels of LIN-3 to ensure the production of 2° cells. This second, relayed signal appears to be a ligand for LIN-12, a Notch-type receptor. Stimulation of LIN-12 on the P5.p and P7.p cells induces expression of a phosphatase that inactivates MAP kinase and affects other regulators as well, thus preventing the 1° fate choice.

In glycogenolysis, signal integration is at the level of a two-subunit protein, with each signal acting on one of the subunits. In vulval cell-fate determination, the activity of a single kinase, MAP kinase, is controlled by two pathways: signaling from an EGF-type receptor activates MAP kinase; signaling from a Notch-type receptor deactivates it. The convergence of these two pathways on MAP kinase elegantly allows the formation of multiple adjacent cell types.

Morphogens Control Cell Fates in Early Drosophila Development

Drosophila has been particularly useful in studying morphogens for three reasons. First, graded regulatory molecules are used extensively in the development of the early Drosophila embryo and in growth of the legs and wings. Second, a fertilized egg develops into an adult fly in only about 10 days. Third, powerful genetic screens have identified many developmental mutants with dramatic abnormal phe-notypes. Some of these defects have been found to arise from mutations in genes encoding morphogens; others arise in genes encoding signal-transduction proteins.

To understand how morphogens determine cell fates in the early fly embryo, we first need to set the scene. Oogenesis begins with a stem cell that divides asymmetrically to generate a single germ cell, which divides four times to generate 16 cells. One of these cells will complete meiosis (see Figure 9-3), becoming an oocyte; the other 15 cells become nurse cells, which synthesize proteins and mRNAs that are transported through cytoplasmic bridges to the oocyte (Figure 15-14). These molecules are necessary for maturation of the oocyte and for the early stages of embryogenesis. At least one-third of the genome is represented in the mRNA contributed by the mother to the oocyte, a substantial dowry. Each group of 16 cells is surrounded by a single layer of somatic cells called the follicle, which deposits the eggshell. The mature oocyte, or egg, is released into the oviduct, where it is fertilized; the fertilized egg, or zygote, is then laid.

The first 13 nuclear divisions of the Drosophila zygote are synchronous and rapid, each division occurring about every 10 minutes. This DNA replication is the most rapid

Nurse cells

Nascent eggshell

Dorsal appendage Egg membrane

Egg shell Polar granules

Follicle (somatic) cells

Nurse cells

Nascent eggshell

Dorsal appendage Egg membrane

Egg shell Polar granules

Follicle (somatic) cells

Dorsal

Anterior —^— Posterior Ventral

Vitelline membrane

Early oogenesis

Mid-oogenesis

▲ FIGURE 15-14 Development of a Drosophila oocyte into a mature egg. A single germ cell gives rise to fifteen nurse cells (green) and a single oocyte (yellow) ( 1|). The early oocyte is about the same size as the neighboring nurse cells; the follicle, a layer of somatic cells, surrounds the oocyte and nurse cells The nurse cells begin to synthesize mRNAs and proteins necessary for oocyte maturation, and the follicle cells begin to form the egg shell. Midway through oogenesis ( 2|), the oocyte has increased in size considerably. The mature egg (3) is surrounded by the completed eggshell (gray). The nurse cells have been discarded,

Mature egg Perivitelline space

Dorsal

Anterior —^— Posterior Ventral

Vitelline membrane

Early oogenesis

Mid-oogenesis

▲ FIGURE 15-14 Development of a Drosophila oocyte into a mature egg. A single germ cell gives rise to fifteen nurse cells (green) and a single oocyte (yellow) ( 1|). The early oocyte is about the same size as the neighboring nurse cells; the follicle, a layer of somatic cells, surrounds the oocyte and nurse cells The nurse cells begin to synthesize mRNAs and proteins necessary for oocyte maturation, and the follicle cells begin to form the egg shell. Midway through oogenesis ( 2|), the oocyte has increased in size considerably. The mature egg (3) is surrounded by the completed eggshell (gray). The nurse cells have been discarded,

Mature egg Perivitelline space but mRNAs synthesized and translocated to the oocyte by the nurse cells function in the early embryo. Polar granules located in the posterior region of the egg cytoplasm mark the region in which germ-line cells will arise. The asymmetry of the mature egg (e.g., the off-center position of the nucleus) sets the stage for the initial cell-fate determination in the embryo. After its release into the oviduct, fertilization of the egg triggers embryogenesis. [Adapted from A. J. F Griffiths et al., 1993, An Introduction to Genetic Analysis, 5th ed., W. H. Freeman and Company, p. 643.]

known for a eukaryote, with the entire 160 Mb of chromosomal DNA copied in a cell-cycle S phase that lasts only 3 minutes. Because these nuclear divisions are not accompanied by cell divisions, they generate a multinucleated egg cell, a syncytium, with a common cytoplasm and plasma membrane (Figure 15-15a). As the nuclei divide, they begin to migrate outward toward the plasma membrane. From about 2 to 3 hours after fertilization, the nuclei reach the surface, forming the syncytial blastoderm; during the next hour or so, cell membranes form around the nuclei, generating the cellular blastoderm, or blastula (Figure 15-15b). All future tissues are derived from the 6000 or so epithelial cells on the surface of the blastula. Soon some of these cells move inside, a process termed gastrulation, and eventually develop into the internal tissues.

The syncytial fly embryo is about 100 cells long, head to tail, and about 60 cells around. Within 1 day of fertilization, the zygote develops into a larva, a segmented form that lacks wings and legs. Development continues through three larval stages (~4 days) and the ~5-day pupal stage during which metamorphosis takes place and adult structures are created (Figure 15-16). At the end of pupation, about 10 days after fertilization, the pupal case splits and an adult fly emerges.

The initially equivalent cells of the syncytial embryo rapidly begin to assume different fates, leading to a well-ordered pattern of distinct cell identities. These early patterning events set the stage for the later development and proper placement of different tissues (e.g., muscle, nerve, epidermis) and body parts, as well as the shapes of the appendages and the organization of cell types within them. Because the early embryo is initially symmetric side to side, the creation of differences among cells is a two-axis prob lem: dorsal/ventral (back/front) and anterior/posterior (head/tail). Different sets of genes act on each axis; so every cell learns its initial fate by responding to input from both dorsoventral-acting and anterioposterior-acting regulators in a kind of two-dimensional grid. As we will see, both regulatory systems begin with information and molecules contributed to the oocyte as a dowry from the mother. When the mature egg is laid, it is already asymmetric along both axes (see Figure 15-14).

Because the early fly embryo is a syncytium, regulatory molecules can move in the common cytoplasm without having to cross plasma membranes. Some molecules form gradients, which are used in the earliest stages of cell-fate determination in Drosophila before subdivision of the syn-cytium into individual cells. Thus transcription factors, as well as secreted molecules, can function as morphogens in the syncytial fly embryo. Syncytia are less common in the early development of other animals and in later stages of fly development; in these stages, patterning events are controlled largely by interactions between cells mediated by extracellular signals, which may act in a graded or relay mode.

To decipher the molecular basis of cell-fate determination and global patterning, investigators have (1) carried out massive genetic screens to identify all the genes having roles in the organizing process, (2) cloned mutation-defined genes; (3) determined the spatial and temporal patterns of mRNA production for each gene and the distribution of the encoded proteins in the embryo; and (4) assessed the effects of mutations on cell differentiation, tissue patterning, and the expression of other regulatory genes. The principles of cell-fate determination and tissue patterning learned from Drosophila have proved to have broad applicability to animal development.

(a) NUCLEAR DIVISION AND MIGRATION

Nuclei with surrounding cytoplasm

(a) NUCLEAR DIVISION AND MIGRATION

Nuclear elongation and extension of cleavage furrows between cells

Cellular blastoderm

(a) Drosophila developmental stages

Nuclear elongation and extension of cleavage furrows between cells

(a) Drosophila developmental stages

Embryonic development

Embryonic development iment 1 day ^

Three larval stages ~4 days iment 1 day ^

Three larval stages ~4 days

Third instar larva

Pupa

DAY 10: Hatching (b) Imaginal discs, precursors to the adult

Third instar larva

Pupa

Cellular blastoderm

DAY 10: Hatching (b) Imaginal discs, precursors to the adult

Mouth part discs Eye-antennal discs Leg discs Wing discs Haltere discs Genital disc

Larva

Mouth part discs Eye-antennal discs Leg discs Wing discs Haltere discs Genital disc

Larva

Adult fly

▲ FIGURE 15-15 Formation of the cellular blastoderm during early Drosophila embryogenesis. Stages from syncytium (a) to cellular blastoderm (b) are illustrated in diagrams and electron micrographs. Nuclear division is not accompanied by cell division until about 6000 nuclei have formed and migrated outward to the plasma membrane. Before cellularization, the embryo displays surface bulges overlying individual nuclei, which remain within a common cytoplasm. No membranes other than that surrounding the entire embryo are present. After cellularization, cell membranes are evident around individual nuclei. Note the segregation of the nuclei of so-called pole cells, which give rise to germ-line cells, at the posterior end of the syncytial blastoderm. [See R. R. Turner and A. P Mahowald, 1976, Devel. Biol. 50:95; photographs courtesy of A. P Mahowald; diagrams after P A. Lawrence, The Making of a Fly, 1992, Blackwell Scientific, Oxford.]

▲ FIGURE 15-16 Major stages in the development of Drosophila. (a) The fertilized egg develops into a blastoderm and undergoes cellularization in a few hours. The larva, a segmented form, appears in about 1 day and passes through three stages (instars) over a 4-day period, developing into a prepupa. Pupation takes ~4-5 days, ending with the emergence of the adult fly from the pupal case. (b) Groups of ectodermal cells called imaginal discs are set aside at specific sites in the larval body cavity. During pupation, these give rise to the various body parts indicated. Other precursor cells give rise to adult muscle, the nervous system, and other internal structures. [Part (a) from M. W. Strickberger, 1985, Genetics, 3d ed., Macmillan, p. 38; reprinted with permission of Macmillan Publishing Company. Part (b) Adapted from same source and J. W. Fristrom et al., 1969, in E. W. Hanly, ed., Park City Symposium on Problems in Biology, University of Utah Press, p. 381.]

Reciprocal Signaling Between the Oocyte and Follicle Cells Establishes Initial Dorsoventral Patterning in Drosophila

Initial dorsal/ventral patterning in Drosophila is controlled by the events of oogenesis. Indeed, the shape of the mature oocyte is an accurate predictor of the dorsal/ventral orientation of the embryo. The process begins when the nucleus of the early oocyte moves slightly, perhaps randomly, toward what will become the anterior and dorsal side of the mature egg (Figure 15-17a). That loss of symmetry triggers the polarization of signals that coordinate the dorsal/ventral axes of the oocyte, embryo, and surrounding eggshell. Such coordination is necessary so that the structures on the eggshell become aligned properly with structures of the growing embryo. For example, breathing tubes in the eggshell must connect with appropriate regions of the embryo.

About midway through Drosophila oogenesis, the production of Gurken, a signal similar to epidermal growth factor, begins. Because of the off-center location of the nucleus,

Gurken is produced on the dorsal side of the oocyte (Figure 15-17b). The receptor for Gurken, a receptor tyrosine kinase like the EGF receptor, is present on the surfaces of all the follicle cells that abut the oocyte. The dorsal Gurken signal activates its receptors only in dorsal follicle cells, leading to changes in their appearance and to repression of the pipe gene within them. Because of this dorsal repression, Pipe protein is produced only in ventral follicle cells. Pipe is an enzyme that catalyzes sulfation of glycosaminoglycans (GAGs), the polysaccharide chains that are added to proteins to form proteoglycans (Chapter 6).

Pipe protein promotes ventral cell fates, probably by activating a still unknown signal that triggers a series of pro-teolytic cleavages in the perivitelline space on the ventral side of the by now mature egg. The ensuing chain of events has some similarity to the blood-clotting cascade, each protein cleaving and thereby activating the next one in the series. The outcome of the cleavages is the production of a ligand called Spätzle only on the ventral side (Figure 15-17c). By this time, the egg has been fertilized and early nuclear division is

(a) Dorsoventral differentiation

(b) Activation of dorsoventral protease cascade

Dorsal follicle cells

Ventral follicle cells

Your Heart and Nutrition

Your Heart and Nutrition

Prevention is better than a cure. Learn how to cherish your heart by taking the necessary means to keep it pumping healthily and steadily through your life.

Get My Free Ebook


Post a comment