developmental exam 2!

Chapter 2. Specification and Patterning

• Developmental biologists study forces within the embryo that cause cells to take on thwir fate as development unforlds. Differentiation- the process producing overt change in appearance or biochemistry in cells. 

How does the fate of a cell become determined? Is it an all or none process? Internal cues in zygitre, egg and early embryo that have cellular interactions and induction- which makes the cells differentiate. 

• As it is committed to a fate, there is restriction of potential fate

  • Specification - labile, will differentiate in neutral environment. They have information about what it can become. 

  • Determination - determined cells will differentiate autonomously even if placed in another region of the embryo. Fate is set and is now irreversible. 

Determined cells are thought to be irreveraibly committed 

• From these levels and types of commitment, different developmental patterns were observed in embryos of different species. 

• Autonomous specification (leads to mosaic developmental pattern)- predetermination, cells from early embryos (blastomeres) will differentiate to their normal fate when isolated 

• This occurs in the correct time frame and with the appropriate cellular structures formed. When the embryo loses these cells, it lacks the structures they would normally have formed. 

• Mosaic developmental patterns seen mostly in invertebrates. 

• Macho development- localized in egg, maternally derived. Has muscle specific actin

• Syncytial specification- one cell, many nuclei, due to incomplete cell division. The DNA replicates without destroying the cyoplasm. 

AUTONOMOUS VS SYNCYTIAL

• Prospective potency = prospective fate. 

• In insects: early developmental patterns due to unequal distribution of cytoplasmic determinants. 

• Bicoid- two tail, caudal

• Nanos gradients make body regions such as head, tail, thorax, abdomen.

• IF SOMETHING DOESNT HAVE A BICOID- THEY COULD HAVE TWO TAILS

• Conditional specification leads to regulative development- cells from early embryos have a flexible fate  

  • Isolation experiment: possibility for twinning- bastomeres can form an entire embryo and can take information from the environment

  • Drieschs demonstration of a regulative developmental pattern in vertebrates and some invertebrates. Pluck blastomeres apart- make more complete embryos. Conditional specification. 

  • Prospective potency > prospective fate. Can indicate conditional specification. Recombination experiment: separate cells so they can mix and reassemble. Reaggregation, Recombination and regression experiment. Transplantation experiment. Defect experiment. Physical pressure to slter nuclei localization. Transfer of cytoplasmic content. 

  • Cellular interactions are important guides- cells direct and restrict each other. 

  • Autonomous specification. Predominant in vertebrates. Invariant cleavages produce the same lineages, blastomere fates are generally invariant. 

WHAT TYPE OF EXPERIMENTS CAN BE DONE TO DETERMINE THE PATTERN AND TIMING OF COMMITMENT FOR A SPECIES? 

• FIND IT, LOSE IT, MOVE IT 

• STRENGTHS OF EVIDENCE: 

  • CORRELATIVE

  • LOSS OF FUNCTION

  • GAIN OF FUNCTION 

  • ACTIN AND MYOCIN ARE HOUSEKEEPING GENES

• FIND IT: FINDING SOMETHING 

• LOSE IT: REMOVE IT, SEE WHAT HAPPENS. IS IT REQUIRED?

• MOVE IT: STICK IT IN DURING A DIFFERENT TIME IN DEVELOPMENT (NEW ENVIRONMENT).

What are sources and how do these cells recieve signals for induction into specific pathways of differentiation to form tissues and organs. 

Current methods for cell analysis of cell identities and fate mapping. 

Chapter 4: Cell- Cell communication and morphogenesis

Early development- whether autonomous or regulative specification. 

• Cleavege to become multicellular

• Regulative specification

• Remodeling and activating zygotic genome

Gastrulation: rearrangement of cells in the blastocyst 

• Patterns of cellular movement

• Cleavage and gastrulation patterns

• Cell adhesion

• Axis specification

How do cells interact for recognition, inductions and development of form? How do cells recognize and react to each other? 

• Reaggregation experiments show that similar cell types adhere to each other

• Types of interactions and components

  • Juxtacrine signaling: hemophilic binding, heteophilic binding, ECM

  • Paracrine Signaling: ligands and receptors 

• Ligands: Proteins that are secreted from a cell and designed to communicate a response in another cell are generally referred to as signaling proteins 

• Receptors: the proteins within a membrane that function to bind either other membrane-associated proteins or signaling proteins 

• Heterophilic binding: adhesion molecules bind to other molecules of the same type

• Hemophilic binding: when adhesion molecules bind to different types

Differentiation: determined fate

Specification: still changeable

Paracrine factors: affect adjacent cells, long distance

Juxtacrine factors: cells must be close to interact

Mesenchymal cells are undifferentiated germ cells. 

Organisms express stage and tissue specific CAMS that arise in cell sorting, ordering and movements dueing morphogenesis. 

• Cell contact and affinity- germ layers cell types and selective affinity evidenced with dissociation and reggregation assays. 

• Quantity and quality of CAMs produces different levels of surface tension and reaggregation, and migration patterns. 

• Cadherens: Calcium dependent adhesion molecules

Cell surface proteins hold cells together and interact with ECM and cytoplasm. CAMS. 

• P- cadherin: Mediates cell-cell adhesion by binding to other P-cadherin molecules on neighboring cells. 

• E- cadherin: a transmembrane protein that plays a crucial role in cell-cell adhesion in epithelial tissues

E-cadherin is widely expressed in most epithelial tissues, while P-cadherin is more restricted to specific basal or myoepithelial cell layers, particularly in the breast tissue, making it considered a marker for these specialized cells; functionally, loss of E-cadherin is often associated with cancer cell invasion, whereas P-cadherin can sometimes play a role in tumor progression depending on the context

• B catenin- structural signaling, links to cytoskeleton

• Integrins- another class of surface adhesion proteins critical for development. FORM DIMERS

• Extracellular- binds to RGD, which is an amino acid sequence that integrins bind to. 

Epithelial mesenchymal transitions – critical for numerous types of morphogenesis. Epithelial cells are cells (from any germ layer) that are found in sheets or tubes, while mesenchymal cells (often mesoderm or neural crest derived) are loosely packed, disconnected cells 

Epithelial cells are able to find each other and look for similar binding sites. 

Extracellular matrices are critical during development

• Proteins: collagen, lamnin and proteoglycans

• Proteoglycans: huge. Many polysaccharides with peptide chains

• Glycoproteins: proteins with sugar groups: fibronectin. 

Induction: the process of one cell type influencing or changing the behavior of another nearby cell

• Requires an inducing cell and a responding cell

• Competence: the ability of the responding cell to respond to inductive signals. 

  • Eye formation in Xenopus: the optic vesicle (part of the neural plate) induces the overlying ectoderm (presumtive epidermis) to form the tissue of the lens

  • Eye formation is due to multiple inductive events. Reciprocal inductions indicate that signaling between cell types continues throughout the differentiation process

Mutations in what types of proteins can cause a failure in these inductive interactions

• Epithelial and mesenchymal interactions: inductive interactions often occur between two general cell types and are common across closely related species. 

  • Epidermal structures of the chick: ther dermis and epidermis induction in the chick shows regional specificity of ther dermal mesenchyme. Depending on the trgion of the origin of the inducting cells, the competent epithelial epidermal cells form different types of structures. 

• Paracrine signaling

  • Local signaling: morphogens

    • Concentration gradients and variable responses TGF-B family and activin as an example

What types of molecules and signaling pathways are involved in induction during development

• FGF and TRK signaling pathways

• Hedgehog pathway

• Cell surfaces with receptors in paracrine signaling. 

Depriving cells of calcium

• Adjacent cells cant find each other 

• Integrins could not bind 

• Calcium binding sites wouldnt work 

• Calcium free media when cells need to not adhere

• EDTA buffer chelating binds to things with 2 charges (calcium, lead, magnesium and heavy metals)

Application: biomaterials for wound healing, 3D tissue printing, cell interaction based on adhesive surfaces

Density and specificity of proteins and glycoproteins and proteoglycans can set up pattern for migration of cells along the ECM. 

• Fibronectin

• Ecm is good for movements in gastrulation 

Timing and adjacency for exposure to signals is important

Axes of bilaterally symmetrical animals 

• Where and when does the information for axis formation arise? Belly vs back 

Case study: 2 heads are better than one

• Proscholdt- how does embryonic axis organize 

• Used DBL cells- DORSAL BLASTOPORE LIP

• Noggin injected into embryo moves signal 

• Chordin and noggin are secreted from the DBL, but do not directly induce dorsal fates. 

• Morphogens can change the fate of tissue- induce it to differentiate via regulatory mechanisms

  • Dorsal vs ventral PH

  • Ventral vs dorsal- gravity 

• Competence: ability to respond to inducing signals. Not passive

• All germ layers are involved in producing

• Competence of ectoderm

• Mutant: change in dna sequence: genome level is called a knockout

Pax6- important in ectoderm- lens formation and competence. Transcription factors

Ingression: sheet to individual 

Integrins: bind to RGD, connects to basal lamina

In order to move away, turn off e- cadherin to degrade in order for it to migrate away and become mesenchymal 

Mesenchyme has regional specificity. Can bind to epithelium to differentiate

induction: 

• Competence is needed as well as an inductive signal

• Multiple and reciprocal inductions

• Often epithelial and mesenchymal interactions

• Key for cellular inyteractions has gastrulation and organogenesis 

Cyclops: 1 eye. Sonic hedgehog gene. 

Chapter 14 and 7

Overarching question: when does human personhood begin? What are key processes that mark development in mammals

Specific stages and processes with homo sapiens, similar to other species, but some differences. 

• Fertilization

• Bastocyst 

• Implantation

• Gastrulation

• Organogenesis

• Fetal growth. Fetus is 9-10 weeks post fertilization

• Birth 

How are the structures of the ganetes optimized for their function 

• Spermatid structure and contents  STRUCTURE. FUNCTION. FATE

  • Tail- motility. Disintegrates 

  • Nucleus- haploid which contains the fathers half of genetic information needed for zygote and development. Sex chromosomes (X or Y). Sperm determines sex. 

  • Enzymes for degradation of ECM (Zona pelluda) found around egg

  • Motochindria: ATP for motility 

  • Cell membrane with specific proteins 

  • Acrosomal vessible- gives reaction for fertilization. 

  • Centiole- organizes motility of tail and becomes centromere, separation of chromosome in mitosis.

• Ovum structure and contents

  • Haploid. Mothers half of genetic information, sex chromosome

  • Cytoplasm for maternal transcripts 

  • ECM for the jelly coat. Importabnt that it sticks around until fertilization. Prevents polyspermy and early implantation

  • Cortical granules- enzymes that prevent polyspermy 

• The ECM, zona pellusa, is important because it has

  • The location where sperm meets egg

  • Helps maintain protective coating

What are potential problems with oocytes or spermatids that may prevent fertilization or development – thus lead to infertility?

• Age. females stop producing eggs (menopause). Males never stop producing but quality diminishes. 

• Dna damage- spermatosis in men

• Disruption of signal molecules- endocrine disruptors in specific pathways

• Endometriosis- female issue with uterine lining

• Immune disorders

• Hormone imbalances- stress and cortisol 

• PCOS- where female makes more testosterone  

• Untreated STD- ectopic pregnancies

• Surgeries

How does IVF overcome some types of infertility (or not) 

• Motility issues such as immotile sperm

• ICSI- IVF

• Ivf is very expensive though- around 20 k 

Infertile: being unable to become preganant for a full year

What mechanisms may be different in internal vs. external fertilization?

• Contact and recognition

• Prevention of polyspermy 

• Fusion of pronuclei

• Activation of egg metabolism 

• Internal vs expernal fertilization

External

• Cues to replicate: 

  • Temp

  • pH

  • Time of year

  • Light

  • Chemically released cues

  • Migration patterns

  • Once one spawns, the others do too. (sea urchins)

  • Number of gametes are higher 

  • Chemoattractants from egg ECM laters, sperm binding and passage through ECM, eventually fusion of egg and sperm membranes

Prevention of polyspermy- 1+ sperm fertilizes an egg- triploid. Extra chromosomes. 

What happens if polyspermy does occur? 

• Triploid

• 4 centromeres

• Double mitosis

• Uneven chromosomes

• Death of embryo

After fertilization- what happens to the successful sperm embryo

• Degrades

• Mitochondira degrades 

• Centroile remains and forms centrosomes

• Sex chromosomes remain

• Nucleus maintains and remodeled 

• Flagellum degrades

• Membrane fuses with egg membrane 

• Nuclear membrane breaks down

During spermatogenesis- dna is packaged with protamines for higher levels of condensation than in somatic cells

Ovum, now zygote, has cytoplasm restructures in the chromatin

• Dna condenses 

• Oocyte derived histones bind and recondense the dna 

• polar body is product of mitosis II

Trophectoderm vs inter cell mass (ICM)

• Trophectoderm becomes 

• ICM becomes part of placenta

Hatching and implantation vs epiblast layer

• Enzymes help degrade ECM

• L selectins make sperm and egg interact. Its a ligand.