cellular mechanisms of development chapter 19. 2 overview of development development is the...

Cellular Mechanismsof Development

Chapter 19

2

Overview of Development

Development is the successive process of systematic gene-directed changes throughout an organism’s life cycle

-Can be divided into four subprocesses:

-Growth (cell division)

-Differentiation

-Pattern formation

-Morphogenesis

3

Cell Division

After fertilization, the diploid zygote undergoes a period of rapid mitotic divisions

-In animals, this period is called cleavage

-Controlled by cyclins and cyclin-dependent kinases (Cdks)

During cleavage, the zygote is divided into smaller & smaller cells called blastomeres

-Moreover, the G1 and G2 phases are shortened or eliminated

4

Cell Division

5

Cell Division

CyclinDegradation

CM

G2

interphase

mitosis

cytokinesis

G2

SG1

M

C

Mitosis

DNA Synthesis

Adult Cell Cycle

S

G1 Cdk /G1cyclin

Cdk /S cyclin

Mitosis

DNA Synthesis

Active

Cell Cycle of Early Frog Blastomere

Active

Active

CyclinSynthesis

Active Cdk /G2

cyclinC

M

SS

M

Cdk /cyclin

Cdk

Inactive

a. b.

6

Cell Division

Caenorhabditis elegans

-One of the best developmental models

-Adult worm consists of 959 somatic cells

-Transparent, so cell division can be followed

-Researchers have mapped out the lineage of all cells derived from the fertilized egg

7

Nematode Lineage Map

a.

b.

Egg andsperm line

Egg

Pharynx

Cuticle-making cells Vulva

Egg Sperm

Adult Nematode

Vulva

GonadGonad

Gonad

Nervous system

Pharynx

Intestine

Cuticle

IntestineNervoussystem

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

8

Cell Division

Blastomeres are nondifferentiated and can give rise to any tissue

Stem cells are set aside and will continue to divide while remaining undifferentiated

-Tissue-specific: can give rise to only one tissue

-Pluripotent: can give rise to multiple different cell types

-Totipotent: can give rise to any cell type

9

Cell Division

Cleave in mammals continues for 5-6 days

producing a ball of cells, the blastocyst

-Consists of:

-Outer layer = Forms the placenta

-Inner cell mass = Forms the embryo

-Source of embryonic stem cells (ES cells)

10

Egg

Sperm Blastocyst Embryo

Embryonic stem-cellculture

Inner cellmass

Once sperm cell and egg cell have joined, cellcleavage produces a blastocyst. The inner cell massof the blastocyst develops into the human embryo.

Embryonic stem cells (ES cells) areisolated from the inner cell mass

11

Cell Division

A plant develops by building its body outward

-Creates new parts from stem cells contained in structures called meristems

-Meristematic stem cells continually divide

-Produce cells that can differentiate into the various plant tissues

-Leaves, roots, branches, and flowers

The plant cell cycle is also regulated by cyclins and cyclin-dependent kinases

12

Cell Differentiation

A human body contains more than 210 major types of differentiated cells

Cell determination commits a cell to a particular developmental pathway

-Can only be “seen” by experiment

-Cells are moved to a different location in the embryo

-If they develop according to their new position, they are not determined

13

Donor No donor

RecipientBefore Overt

Differentiation

RecipientAfter Overt

Differentiation

Normal Not Determined(early development)

Determined(later development)

Tail cells aretransplanted

to head

Tail cells developinto head cells in head

Tail cells developinto tail cells in head

Tail cells aretransplantedto head

Tail Head


14


Cells initiate developmental changes by using transcriptional factors to change patterns of gene expression

Cells become committed to follow a particular developmental pathway in one of two ways:

1) via differential inheritance of cytoplasmic determinants

2) via cell-cell interactions

15


Cytoplasmic determinants

-Tunicates are marine invertebrates

-Tadpoles have tails, which are lost during metamorphosis into the adult

-Egg contains yellow pigment granules

-Become asymmetrically localized following fertilization

-Cells that inherit them form muscles

16


a.

n

2n

Embryo(diploid) 2n

Larva(diploid) 2n

Sperm(haploid) n

Egg(haploid) n

Pigmentgranules

Adult tunicate(diploid) 2n

ME

IOS

IS

MEIOSIS

METAMORPHOSIS

17


Cytoplasmic determinants

-Female parent provides egg with macho-1 mRNA

-Encodes a transcription factor that can activate expression of muscle- specific genes

18


Induction is the change in the fate of a cell due to interaction with an adjacent cell

If cells of a frog embryo are separated:

-One pole (“animal pole”) forms ectoderm

-Other pole (“vegetal pole”) forms endoderm

-No mesoderm is formed

If the two pole cells are placed side-by-side, some animal-pole cells form the mesoderm

19


Another example of induction is the formation of notochord and mesenchyme in tunicates

-Arise from mesodermal cells that form at the vegetal margin of 32-cell stage embryo

-Cells receive a chemical signal from underlying endodermal cells

-Anterior cells differentiate into notochord

-Posterior cells differentiate into mesenchyme

20

Anterior

Posterior

Anterior Posterior

a.

b. c.

21

FGF signaling

Sagittal section Longitudinal section

Longitudinal section

Dorsal nerve cord (NC)

Mesenchymal cells (Mes)

Tail muscle cells (Mus)

32-Cell Stage 64-Cell Stage

Anterior

Posterior

2

1

Notochord (Not)Ventral endoderm (En)


21


The chemical signal is a fibroblast growth factor (FGF) molecule

-The FGF receptor is a tyrosine kinase that activates a MAP kinase cascade

-Produces a transcription factor that triggers differentiation

Thus, the combination of macho-1 and FGF signaling leads to four different cell types

22


a.

FGF Signal received?Mesenchyme

MuscleNotochord

FGF Signal received?

Macho-1 inherited?

Nerve cord

First Step Cell TypesSecond Step

YesNo

YesNo

Yes

No

FGF

FGF Receptor

Ras/MAPKPathway

T-Ets Macho-1

Cell membrane

P

MesenchymePrecursor Cells

Suppression of musclegenes and activation

of mesenchyme genes

b.

23

Cell DifferentiationNo

FGFFGF Receptor

Ras/MAPKPathway

T-Ets

Cell membrane

Nerve cordPrecursor Cells

FGF

FGF Receptor

Transcriptionof notochord genes

P

Notochord Precursor Cells

NoFGF

FGF Receptor

Ras/MAPKPathway

Transcription of muscle genes

MusclePrecursor Cells

Suppression of notochordgenes and activationof nerve cord genes

No Macho-1No Macho-1Macho-1

Cell membraneCell membrane

T-EtsT-Ets

Ras/MAPKPathway

24

Cloning

Until very recently, biologists thought that determination and cell differentiation were irreversible in animals

Nuclear transplant experiments in mammals were attempted without success

-Finally, in 1996 a breakthrough

Geneticists at the Roslin Institute in Scotland performed the following procedure:

25

Cloning

1. Differentiated mammary cells were removed from the udder of a six-year old sheep

2. Eggs obtained from a ewe were enucleated

3. Cells were synchronized to a resting state

4. The mammary and egg cells were combined by somatic cell nuclear transfer (SCNT)

5. Successful embryos (29/277) were placed in surrogate mother sheep

6. On July 5, 1996, Dolly was born

26

Development Implantation Birth of Clone Growth to Adulthood

Embryo begins todevelop in vitro.

Embryo isimplanted into

surrogatemother.

After a five-month pregnancy, alamb genetically identical to thesheep from which the mammarycell was extracted is born.

Embryo

Preparation Cell Fusion Cell Division

Mammary cell is extracted and grown in nutrient-deficient solution that arrests the cell cycle.

Egg cell is extracted. Nucleus is removed fromegg cell with a micropipette.

Nucleus containingsource

Mammary cell isinserted inside

covering of egg cell.

Electric shock fuses cellmembranes and triggers

cell division.

27

Cloning

Dolly proved that determination in animals is reversible

-Nucleus of a differentiated cell can be reprogrammed to be totipotent

Reproductive cloning refers to the use of SCNT to create an animal that is genetically identical to another

-Scientists have cloned cats, rabbits, rats, mice, goats and pigs

28

Cloning

Reproductive cloning has inherent problems

1. Low success rate

2. Age-associated diseases

Normal mammalian development requires precise genomic imprinting

-The differential expression of genes based on parental origin

Cloning fails because there is not enough time to reprogram the genome properly

29

Cloning

In therapeutic cloning, stem cells are cloned from a person’s own tissues and so the body readily accepts them

Initial stages are the same as those of reproductive cloning

-Embryo is broken apart and its embryonic stem cells extracted

-Grown in culture and then used to replace diseased or injured tissue

30

Cloning

The skin cellnucleus is insertedinto the enucleated

human egg cell.

Cell cleavageoccurs as the

embryo begins todevelop in vitro.

The embryoreaches the

blastocyst stage

The nucleus from a skin cell of a diabeticpatient is removed.

The nucleus from a skin cell of a healthy patient is removed.

Early embryo Blastocyst

Inner cellmass

EScells

Diabeticpatient

Healthypatient

31

CloningTherapeutic Cloning

Reproductive Cloning

Embryonic stem cells(ES cells) are extractedand grown in culture.

The blastocyst is kept intact andis implanted into the uterus of a

surrogate mother.

The resulting baby isa clone of the

healthy patient.

The stem cells are developedinto healthy pancreatic islet cells

needed by the patient.

The healthy tissue isinjected or transplantedinto the diabetic patient.

Healthy pancreatic islet cells

Diabeticpatient

32

Cloning

Human embryonic stem cells have enormous promise for treating a wide range of diseases

-However, stem cell research has raised profound ethical issues

Very few countries have permissive policy towards human reproductive cloning

-However, many permit embryonic stem cell research

33

Cloning

Early reports on a variety of adult stem cells indicated that they may be pluripotent

-Since then these results have been challenged

34

Pattern Formation

In the early stages of pattern formation, two perpendicular axes are established

-Anterior/posterior (A/P, head-to-tail) axis

-Dorsal/ventral (D/V, back-to-front) axis

Polarity refers to the acquisition of axial differences in developing structures

Position information leads to changes in gene activity, and thus cells adopt a fate appropriate for their location

35

Drosophila Embryogenesis

Drosophila produces two body forms

-Larva – Tubular eating machine

-Adult – Flying sex machine axes are established

Metamorphosis is the passage from one body form to another

Embryogenesis is the formation of a larva from a fertilized egg

36


Before fertilization, specialized nurse cells move maternal mRNAs into maturing oocyte

-These mRNA will initiate a cascade of gene activations following fertilization

Embryonic nuclei do not begin to function until approximately 10 nuclear divisions later

37


After fertilization, 12 rounds of nuclear division without cytokinesis produces a syncytial blastoderm

-4000 nuclei in a single cytoplasm

Membranes grow between the nuclei forming the cellular blastoderm

Within a day of fertilization, a segmented, tubular body is formed

38

c.

b.

Nursecells

AnteriorPosterior

Movement ofmaternal mRNA

Oocyte

Folliclecells

Fertilized egg

a.

d.

e.

Three larval stages

Syncytial blastoderm

Cellular blastoderm

Nuclei line up alongsurface, and membranesgrow between them toform a cellular blastoderm.

Segmented embryo prior to hatching

Metamorphosis

Abdomen

Thorax

Head

Hatching larva


Nucleus

Embryo

39


Nüsslein-Volhard and Wieschaus elucidated how the segmentation pattern is formed

-Earned the 1995 Nobel Prize

Two different genetic pathways control the establishment of the A/P and D/V polarity

-Both involve gradients of morphogens

-Soluble signal molecules that can specify different cell fates along an axis

40

About 21/2 hours after fertilization, bicoid protein turns on a series of brief signals from so-called gap genes. The gap proteins act to divide the embryo into large blocks. In this photo, fluorescent dyes in antibodies that bind to the gap proteins Krüppel (orange) and Hunchback (green) make the blocks visible; the region of overlap is yellow.

Forming the SegmentsLaying Down the Fundamental Regions

Setting the Stage for Segmentation

Bicoid

Hairy

Krüppel Hunchback

Engrailed

Establishing the Polarity of the Embryo


Fertilization of the egg triggers the production of bicoid protein from maternal RNA in the egg. The bicoid protein diffuses through the egg, forming a gradient. This gradient determines the polarity of the embryo, with the head and thorax developing in the zone of high concentration (green fluorescent dye in antibodies that bind bicoid protein allows visualization of the gradient).

About 0.5 hr later, the gap genes switch on the “pair-rule” genes, which are each expressed in seven stripes. This is shown for the pair-rule gene hairy . Some pair-rule genes are only required for even-numbered segments while others are only required for odd numbered segments.

The final stage of segmentation occurs when a “segment- polarity” gene called engrailed divides each of the seven regions into halves, producing 14 narrow compartments. Each compartment corresponds to one segment of the future body. There are three head segments (H, bottom right), three thoracic segments (T, upper right), and eight abdominal segments (A, from top right to bottom left).

41

Establishment of the A/P axis

Nurse cells secrete maternally produced bicoid and nanos mRNAs into the oocyte

-Differentially transported by microtubules to opposite poles of the oocyte

-bicoid mRNA to the future anterior pole

-nanos mRNA to the future posterior pole

-After fertilization, translation will create opposing gradients of Bicoid and Nanos proteins

42

a.

b.

bicoid mRNA movestoward anterior end

bicoidmRNA

Movement ofmaternal mRNA

Nucleus

Microtubules

Nursecells

Folliclecells

nanos mRNA movestoward posterior end

PosteriorAnterior

PosteriorAnterior

nanosmRNA


43

Establishment of the A/P axis

Bicoid and Nanos control translation of two other maternal mRNAs, hunchback and caudal, that encode transcription factors

-Hunchback activates anterior structures

-Caudal activates posterior structures

The two mRNAs are not evenly distributed

-Bicoid inhibits caudal mRNA translation

-Nanos inhibits hunchback mRNA translation

44

Co

nce

ntr

atio

n

Anterior

Anterior

a. Oocyte mRNAs

c. Early cleavage embryo proteins

b. After fertilization

Posterior

Posterior

nanos mRNA

hunchback mRNA

bicoid mRNA

caudal mRNA

Nanos protein

Hunchback protein

Bicoid protein

Caudal proteinC

on

cen

trat

ion

Anterior Posterior

nanos mRNAhunchback mRNAbicoid mRNAcaudal mRNA

Nanos proteinHunchback proteinBicoid proteinCaudal protein


45

Establishment of the D/V axis

Maternally produced dorsal mRNA is placed into the oocyte

-Not asymmetrically localized

Oocyte nucleus synthesizes gurken mRNA

-Accumulates in a crescent on the future dorsal side of embryo

After fertilization, a series of steps results in selected transport of Dorsal into ventral nuclei, thus forming a D/V gradient

47

Production of Body Plan

The body plan is produced by sequential activation of three classes of segmentation genes

1. Gap genes

-Map out the coarsest subdivision along the A/P axis

-All 9 genes encode transcription factors that activate the next gene class

48


2. Pair-rule genes

-Divide the embryo into seven zones

-The 8 or more genes encode transcription factors that regulate each

other, and activate the next gene class

3. Segment polarity genes

-Finish defining the embryonic segments

49


Segment identity arises from the action of homeotic genes

-Mutations in them lead to the appearance of normal body parts in unusual places

-Ultrabithorax mutants produce an extra pair of wings

50


Homeotic gene complexes

-The HOM complex genes of Drosophila are grouped into two clusters

-Antennapedia complex, which governs the anterior end of the fly

-Bithorax complex, which governs the posterior end of the fly

-Interestingly, the order of genes mirrors the order of the body parts they control

51


Homeotic gene complexes

-All of these genes contain a conserved 180-base sequence, the homeobox

-Encodes a 60-amino acid DNA-binding domain, the homeodomain

-Homeobox-containing genes are termed Hox genes

-Vertebrates have 4 Hox gene clusters

52


Drosophila HOM genes

Thorax

Antennapedia complex

Head Abdomen

Bithorax complex

Fruit fly

Fruit flyembryo

Mouse

Hox 1

Hox 2

Hox 3

Hox 4

Mouseembryo

a. b.

Drosophila HOM Chromosomes Mouse Hox Chromosomes

lab pb Dfd Scr Antp Ubxabd-Aabd-B

53

Pattern Formation in Plants

The predominant homeotic gene family in plants is the MADS-box genes

-Found in most eukaryotic organisms, although in much higher numbers in plants

MADS-box genes encode transcriptional regulators, which control various processes:

-Transition from vegetative to reproductive growth, root development and floral organ identity

54

Morphogenesis

Morphogenesis is the formation of ordered form and structure

-Animals achieve it through changes in:

-Cell division

-Cell shape and size

-Cell death

-Cell migration

-Plants use these except for cell migration

55

Morphogenesis

Cell division

-The orientation of the mitotic spindle determines the plane of cell division in eukaryotic cells

-If spindle is centrally located, two equal-sized daughter cells will result

-If spindle is off to one side, two unequal daughter cells will result

56

Morphogenesis

Cell shape and size

-In animals, cell differentiation is accomplished by profound changes in cell size and shape

-Nerve cells develop long processes called axons

-Skeletal muscles cells are large and multinucleated

57

Morphogenesis

Cell death

-Necrosis is accidental cell death

-Apoptosis is programmed cell death

-Is required for normal development in all animals

-“Death program” pathway consists of:

-Activator, inhibitor and apoptotic protease

58

a. b.

InhibitorCED-9 Bcl-2

CED-4 Apaf1

Caspase-8 or -9

Apoptosis

CED-3

Apoptosis

Inhibitor:

Activator:

ApoptoticProtease:

Caenorhabditis elegans Mammalian CellOrganism

Inhibition

Activation


59

Morphogenesis

Cell migration

-Cell movement involves both adhesion and loss of adhesion between cells and substrate

-Cell-to-cell interactions are often mediated through cadherins

-Cell-to-substrate interactions often involve complexes between integrins and the extracellular matrix (ECM)

60

Development of Seed Plants

Plant development occurs in five main stages:

1. Early embryonic cell division

-First division is off-center

-Smaller cell divides to form the embryo

-Larger cell divides to form suspensor -Cells near it ultimately form the root -Cells on the other end, form the

shoot

61


2. Embryonic tissue formation-Three basic tissues differentiate:

-Epidermal, ground and vascular3. Seed formation

-1-2 cotyledons form-Development is arrested

4. Seed germination-Development resumes -Roots extend down, and shoots up

62


5. Meristematic development and morphogenesis

-Apical meristems at the root and shoot tips generate a large numbers of cells

-Form leaves, flowers and all other components of the mature plant

63

a. Early cell division

Embryo

Embryo

Suspensor

b. Tissue formation

d. Germinationc. Seed formation meristem

Seed wall

Shoot apicalmeristem

Root apical

Shoot apical meristem

Cotyledons

e. Meristematic development and morphogenesis

Root apicalmeristem

Epidermalcells

Groundtissue cellsVasculartissue cells

Cotyledons


64

Environmental Effects

Both plant and animal development are affected by environmental factors

-Germination of a dormant seed proceeds only under favorable soil and day conditions

-Reptiles have a temperature-dependent sex determination (TSD) mechanism

-The water flea Daphnia changes its shape after encountering a predatory fly larva

65


66


In mammals, embryonic and fetal development have a longer time course

-Thus they are more subject to the effects of environmental contaminants, and blood-borne agents in the mother

-Thalidomide, a sedative drug

-Many pregnant women who took it had children with limb defects

67


Endocrine disrupting chemicals (EDCs)

-Interfere with synthesis, transport or receptor-binding of endogenous hormones

-Derived from three main sources

-Industrial wastes (polychlorinated biphenyls or PCBs)

-Agricultural practices (DDT)

-Effluent of sewage-treatment plants

cellular mechanisms of development chapter 19. 2 overview of development development is the...

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