Developmental Biology Lecture #6

Early Embryonic Development

• Steps after Fertilization:

  1. Cleavage: Fast division of cells.

  2. Gastrulation: Movement of cells with respect to each other, forming germ layers.

  • Axis Formation: Necessary to establish three axes:

  1. Anterior to posterior

  2. Dorsal to ventral

  3. Left to right

Introduction to Cleavage

• Overview of Cleavage:

  • Egg Division: The egg divides into many smaller cells termed blastomeres.

  • Maternal Influence: The rate of division and placement of blastomeres is primarily controlled by maternal mRNA.

  • Nuclear Division: This phase exhibits a rapid nuclear division rate, unparalleled at other stages.

  • Cell Growth: Cells do not grow between divisions; there is no G1 or G2 phase present.

Cyclin Regulation of the Cell Cycle

• Role of Cyclins:

  • Cell Cycle Regulation: Cyclins play a critical role in regulating steps of the cell cycle.

  • Mitosis-Promoting Factor (MPF):

  • Composed of cyclin B and cyclin-dependent kinase.

  • Accumulates during M phase and is degraded during S phase.

  • Functions:

    • Phosphorylates histones

    • Phosphorylates nuclear envelope lamin

    • Regulates myosin leading to chromatin condensation

    • Affects nuclear envelope depolymerization and formation of mitotic spindle

Mid-Blastula Transition (MBT)

• Progression:

  • Stored Proteins: Cyclin B is regulated by stored proteins, and as cytoplasmic components are utilized, the nucleus commences synthesizing mRNA.

  • Initiation of MBT:

  • Embryo starts to make its own mRNA.

  • The G1 and G2 phases are incorporated into the cell cycle.

  • Cell cycles thereafter are no longer synchronized.

Cleavage Patterns

• Influencing Factors:

  • Yolk Placement: Determines cleavage patterns between different embryos.

  • Inherited Factors: Influence the angle of the mitotic spindle.

• Division Poles:

  • Embryo divided into animal and vegetal poles.

  • More yolk accumulates in the vegetal pole, while the nucleus is positioned toward the animal pole.

Types of Cleavage Patterns

I. Holoblastic Cleavage

• A. Isolecithal Cleavage:

  1. Radial Cleavage: Examples include Echinoderms and amphioxus.

  2. Spiral Cleavage: Found in annelids, molluscs, and flatworms.

  3. Bilateral Cleavage: Seen in tunicates.

  4. Rotational Cleavage: Observed in mammals and nematodes.

• B. Mesolecithal Cleavage:

  • Displaced Radial Cleavage: Notable in amphibians.

II. Meroblastic Cleavage

• A. Telolecithal Cleavage:

  1. Bilateral Cleavage: Seen in cephalopod molluscs.

  2. Discoidal Cleavage: Observed in fish, reptiles, and birds.

• B. Centrolecithal Cleavage:

  • Superficial Cleavage: Characteristic of most insects.

Gastrulation

• Definition: Establishment of a multilayered body plan, which leads to three distinct layers:

  • Ectoderm

  • Mesoderm

  • Endoderm

• Types of Gastrulation Movements:

  • Complex movements categorized into five primary types.

Axis Formation in the Embryo

• Three Axes Formed:

  1. Anterior to posterior axis

  2. Dorsal to ventral axis

  3. Left-right axis

Early Development in Drosophila

• Importance of Drosophila:

  • First organism extensively studied in genetics due to ease of breeding and identifying mutants.

  • Challenges in examining early embryology due to small size and difficulty in transplantation.

  • Advances in molecular biology have transformed the study of early development.

Embryology in Drosophila

• Cellular Dynamics:

  • The early embryo is a single cell with multiple nuclei until the 13th division.

  • Nuclei migrate to the outer edge of the embryo, establishing polarity determined by placement within the ovary and interaction with follicle cells.

• Cleavage Process:

  • Superficial Cleavage: The outer edge becomes cellularized while the middle retains yolk.

  • Nuclei divide in the center and migrate to the periphery, forming pole cells that become gametes.

  • Initially, nuclei have no membranes between them, leading to a syncytial blastoderm that cellularizes at the 13th cycle to form a cellular blastoderm.

Mid-Blastula Transition in Drosophila

• Transition Details:

  • Nuclear division slows once nuclei reach the periphery of the embryo.

  • Division becomes asynchronous upon cellularization.

  • Nuclear transcription begins around the 11th cycle, signaling the mid-blastula transition.

Gastrulation in Drosophila

• Initiation of Gastrulation:

  • Begins at mid-blastula transition with mesoderm invaginating to form the ventral furrow, which later pinches off to create a tube that covers the ectoderm.

  • Endoderm invaginates anterior and posterior to the ventral furrow, creating two tubes that eventually merge, including internalization of pole cells with the endoderm.

Developmental Gradients in Drosophila

• Establishment of Anterior to Posterior Axis:

  • Classic embryology suggests gradients shape development.

  • Experiments show that tying off one end of the embryo yields either anterior or posterior portions.

  • Research by Christiane Nusslein-Volhard identified mutations in early patterning genes leading to a Nobel Prize in 1995.

• Gradient Generators:

  • Gradients established by proteins such as bicoid and hunchback (anterior) and nanos and caudal (posterior), which are crucial in forming abdominal segments.

  • Bicoid is tethered to microtubules at the anterior, while nanos binds to the cytoskeleton in the posterior.

  • Hunchback and caudal are distributed throughout the embryo.

Setting Up Gradients and Gene Activation

• Role of Nurse Cells:

  • Microtubules oriented by nurse cells direct mRNA locations within the egg.

  • mRNA binds to proteins (dynein or kinesin) that transport them to opposite ends of the embryo.

• Gradient Influence on Gene Activation:

  • Initial gradients activate gap genes, the first genes to be transcribed.

  • Variations in gap gene concentrations activate pair-rule genes, leading to series of developmental segments.

Segmentation Genes in Drosophila

• Classification of Segmentation Genes:

  • Gap Genes: Activated by maternal gene gradients. Mutations in gap genes cause gaps in anatomical structures.

  • Pair-rule Genes: Activated by gap genes, dividing the embryo into parasegments with a striped pattern. Enhancers for pair-rule genes respond variably to different concentrations of gap gene products.

  • Segment Polarity Genes: Maintain structure's periodicity and are expressed in cellularized embryos. These genes require intercellular communication for function.

Homeotic Selector Genes

• Function and Identification:

  • After segment boundaries are established, homeotic selector genes are utilized to identify segments, arranged in an anterior to posterior order.

  • These genes dictate the fate of segments in development.

• Homeotic Mutations:

  • Loss of homeotic selectors leads to misplacements or duplications of body structures, e.g., deletion of ultrabithorax causes two second thoracic segments, while misexpression of Antennapedia leads to leg growth from the head.

• Stabilization of Homeotic Gene Expression:

  • Homeotic gene expression becomes permanent, with repression by posterior genes.

  • The transcription of these genes is assimilated into chromatin, with active chromatin configurations maintained following initial activation.

Dorsal-Ventral Axis Formation

• Dorsal Protein Contribution:

  • Concentrations of Dorsal protein establish the ventral versus dorsal identities.

  • Dorsal protein is produced throughout the embryo but only enters nuclei on the ventral side; in other regions, it is retained in the cytoplasm.

• Mechanism of Dorsal Entry:

  • The oocyte nucleus's location signals follicle cells, directing the dorsal side.

  • Follicle cells lacking a signal form the ventral region.

Experimental Embryology in Amphibians

• Organism Selection:

  • Amphibians, particularly useful due to large oocyte size and rapid development.

  • Challenges include prolonged maturation times and organisms having tetraploid (4 copies) genomes.

  • New molecular techniques are assisting in genetic determination.

Cleavage in Amphibians

• Embryonic Characteristics:

  • Radially symmetric and holoblastic embryos with significant yolk in the vegetal hemisphere.

  • Cleavage furrows progress slowly into the vegetal region, regulated by mitosis-promoting factors (MPF).

• Blastocoel Formation:

  • At the 128-cell stage, a blastocoel forms, serving two functions:

  1. Creating necessary space for gastrulation movements.

  2. Separating vegetal cells from inducing influences on animal cells.

Xenopus Fate Mapping

• Mapping Overview:

  • The mapped structures include:

  • Epidermis

  • Supra-blastoporal endoderm

  • Subblastoporal endoderm

  • Neural plate, lateral plate mesoderm

  • Somites and notochord

  • Blastopore

  • Heart

  • The pattern is established in unfertilized eggs through factors like VegT transcription factor and TGF-B paracrine factor Vg1, with depletion of VegT leading to the absence of endoderm and limited mesoderm.

Cell Movements During Gastrulation

• Blastopore Formation:

  • The blastopore, a future dorsal side, forms where animal and vegetal regions meet.

  • Cell Types: Bottle cells enter first, forming the archenteron which begins to displace the blastocoel.

• Cell Involution and Epiboly:

  • Involution: Marginal zone cells undergo involution while animal cells undergo epiboly as they reach the dorsal blastopore lip.

  • Endoderm and mesoderm involute at the ventral blastopore lip.

• Final Structure Formation:

  • The yolk plug forms and eventually the blastopore closes, surrounding the embryo with ectoderm.

Mid-Blastula Transition and Gastrulation

• Triggering Events of MBT:

  • Occurs at the 12th cell cycle with the beginning of nuclear transcription.

  • Initiated by demethylation of specific promoters, permitting transcription factors to bind and activate targeted genes.