Polarity and Segmentation. Chapter Two

Transcription

1 Polarity and Segmentation Chapter Two

2 Polarization Entire body plan is polarized One end is different than the other Head vs. Tail Anterior vs. Posterior Front vs. Back Ventral vs. Dorsal Majority of neural tube = spinal cord Anterior end = brain

3 Vertebrate body is symmetrical but polarized (tail vs. head) How does polarization of the animal body originate during development?

4 In vertebrates, the rostral portion of neural tube gives origin to brain structures Caudal (tail) areas of neural tube give origin to spinal cord

5 CNS Subdivided CNS = central nervous system Spinal cord Derived from neural tube directly Brain Specialized anterior section of neural tube Brain has three primary divisions: Prosencephalon forebrain Mesencephalon midbrain Rhombencephalon hindbrain

6 3 primary brain vesicles are further divided: Prosencephalon 1. Telencephalon: Cerebral hemispheres 2. Diencephalon: Thalamus, hypothalamus, and optic vesicles Mesencephalon remains Rhombencephalon 1. Metencephalon Cerebellum 2. Myelencephalon Medulla

7 In Drosophila, development of the anterior portion of the nervous system also undergo a three-chamber pattern: 1) Protocerebrum 2) Deutocerebrum 3) Tritocerebrum The main difference here is that the ventral nerve cord is generated by delamination of epithelial cells that are fused together into interconnected ganglia So how do regional differences between rostral and caudal structures originate in vertebrates and insects?

8 Formation of the Anterior-Posterior Axis Anterior In Drosophila Posterior In Drosophila, the A-P axis is set by two molecules: 1) bicoid 2) nanos Anterior Posterior These two molecules create an opposing gradient Opposing gradient separates Anterior from Posterior Triggers other genes to be on or off

9 A/P axis in Drosophila Levels of development: 1. Cytoplasm gradients of nanos and bicoid 1. Inherited from mother s oocyte 2. Gap genes 3. Pair rule genes 4. Segment polarity genes 5. Homeotic genes 1. Trigger regional gene expression differences

10 A/P axis in Drosophila

11 Formation of the Anterior-Posterior Axis In Drosophila Anterior Posterior Normal development of anterior region in nanos -/- Lack of anterior region in bicoid mutant Development of anterior structures following injection of bicoid protein in posterior region Exact reverse for Nanos and posterior development

12 Once the A-P layout is defined, what factors determine the differentiation of each body segment? Once A/P axis is laid out Now each segment must express unique genes in order to be different In Drosophila, differentiation of each segment requires the expression of Hox homeobox genes Homeobox genes are all transcription factors

13 Once the A-P layout is defined, what factors determine the differentiation of each body segment? Homeobox genes are arranged in a linear array on the chromosome Homeobox genes at the 3 end are expressed in more anterior locations Homeobox genes control regional identity of body segment

14 Once the A-P layout is defined, what factors determine the differentiation of each body segment? Homeobox genes are conserved among animal species Vertebrates: Have more hox genes Complex interactions Overlap of same function Still exist in order 3 to 5

15 Homeobox genes Hox genes Always transcription factors Bind DNA directly: Through the homeobox domain Activate the genes that directly cause specific regional identity Deactivate other genes

16 Distaless is NOT a homeobox gene Insects have three pairs of legs One pair on each thoracic segment No legs on the abdominal segments Distaless gene forms the legs Distaless expression is suppressed in abdominal segments of insect By BT-X a hox gene BT-X binds the DNA and repressed distaless

17 Example of Hox mutations: wild type Antennapedia Mutation in the antennapedia complex result in the formation of a leg where an antenna was supposed to exist

18 Do Hox genes control vertebrate development as well? XlHbox1 in Xenopus Regional expression pattern of XlHbox1

19 Do Hox genes control vertebrate development as well? Injection of an antibody against the XlHbox1 protein result in the enlargement of the hindbrain

20 Knock Out Mice Specific genes deletion studies Study the role of homeobox genes in segmental differentiation

21 Hox knockouts have allowed the study of regional differences in the developing mice hindbrain Segmentation of the hindbrain result in the formation of rhombomeres similar to Drosophila s segments Rhombomere formation in the mouse is encoded by different combination of Hox homeobox genes. Hox mutations affect the development of specific rhombomeres

22 Hox knockouts have allowed the study of regional differences in the developing mice hindbrain Deletion of Hoxa1 gene results in fusion of rhombomeres (R) 5 & 6 and reduction of R4 Deletion of Hoxb1 gene results the loss of motoneurons R5 Double mutants of Hoxb1 & Hoxa1 genes show a combined effect

23 Segmental Specification is Encoded by Combination of Factors Segmental arrangement of the chick hindbrain rhombomeres (r1-r7) result in a steroptypic pattern of motoneuron localization in hindbrain Early transcription factors, Eph family receptors, and homeobox genes establish the segmental specification of the hindbrain rhombomeres

24 Important to Note Vertebrate hindbrain segmentation Occurs by exact same mechanism as insect body segmentation Express different hox genes Produce different regional identities In insects get body segments In vertebrates get different motor neurons In exactly to the correct brain region

25

26 Removing Hox genes: Entire hindbrain looks like R1 segment

27 What controls Hox genes? Step backwards one If Hox genes regulate all other genes What regulates Hox genes? In flies: Cytoplasmic gradients Control Gap genes Control Pair rule genes control Hox genes Same in vertebrates?

28 What signal molecules pattern the Hox expression? Retinoic acid (RA) has been show to regulate Hox gene expression. RA crosses cell membrane and bind to cytoplasmic receptors. The RA-receptor complex can translocate to the nucleus and regulate gene expression after binding to RA response element (RARE)

29 There is a gradient of RA expression in the developing embryo RA levels are significantly higher in posterior regions of Xenopus embryos RA normally activates posterior identity Suppresses anterior identity Exposure of the developing embryo to RA results in malformations of anterior structures (head structures fail to develop)

30 Retinoid Acid Controls Formation of A-P Axis and Hox expression in vitro and in vivo Low RA results in Hox genes normally expressed in the anterior portion High RA results in Hox genes normally expressed in the posterior portion of the embryo Target deletion of RA receptors results in head structure formation RA normally regulates in more posterior regions

31 Heads vs. Tails? Spemann and others proved there were both head and tail organizers This meant that if you transplanted a small piece of tissue from head to anywhere it would still form into a head Same with piece of tail tissue What are in these regions? Transcription factors that induce other genes

32 Heads vs. Tails? Nieuwkoop transplanted small piece of head tissue to different places along axis All pieces transformed into head tissue However, if transplanted into caudal regions actually formed two tissue types: Anterior and Posterior both developed New theory: Activator = 1 st signal Transformer = 2 nd signal

33 Activator-Transformer Hypothesis Activator = gene that turns ectodermal cells into neural tissue Anterior is default state of neural tissue Transformer = gene that turns neural tissue into posterior types Posterior requires two signals: Neural positive Posterior positive

34 Activator Genes Genes that induce neural tissue Noggin, Chordin, Follistatin All produce anterior structures Therefore anterior must be default for neural tissue Remove these activator signals and get NO neural tissue Rather than posterior-like tissue

35 Transformer Genes Retinoic acid Posteriorize embryos Regulates hindbrain hox genes Wnt and beta-catenin When wnt is inhibited a second head can form Adding wnt posteriorizes neural tissue FGF Induce posterior gene expression

36 The Activator-Transformer Hypothesis Blocking of BMP signal enables formation of neural tissue. In this case noggin, chordin, follistatin play the role of neural inducers that allow formation of anterior structures Formation of a retinoid acidgradient enables polarization of the neural tissue. In this case RA is the transformer that promotes Hox gene expression and formation of caudal structures

37 Complexity Although anterior type seems to be default Suppressing BMPs is necessary to form anterior tissue Suppressing BMPs is NOT sufficient to form functional anterior tissue If all you do is suppress BMPs you will form anterior structures Will not be fully functional or normal

38 Anterior Tissues Anterior tissues require two signals: 1. Inhibition of BMPs 1. Forms neural tissue 2. Inhibition of wnt pathway 1. Anteriorizes the neural tissue completely Inhibition of BMPs is necessary, but not sufficient to form anterior structures Inhibition of wnt pathway is necessary but not sufficient

39 Role of wnt in Xenopus embryos G = Suppressing BMPs and wnt pathways I = Suppressing BMPs alone Small head and cyclopia

40 Wnt inhibitors All expressed within organizer: Cerberus Injection of Cerberus will cause ectopic head formation frzb Injection forms larger than normal heads dkk1 Ectopic head formation

41 Wnt Remove wnt = extra head Inhibit wnt = extra head Remove wnt inhibitor = messed up head

42 Cooperation BMP inhibitors work together with wnt inhibitors to form head and brain Wild Type Double mutant: dkk and Noggin

43 FGF Third transformer is FGF Both FGF and wnt signals suppress expression of enzyme cyp26 cyp26 is an enzyme that breaks down retinoic acid Without cyp26 RA builds up: Posteriorization of tissues Exact mechanism/interaction of FGF and wnt is unknown

44 Any Questions? Read Chapter Two