1. Looking for the appropriate size: genetics under control Looking for the appropriate size: genetics under control Crazy about Biomedicine– May 2013 Ana Ferreira Development and Growth Control Lab

2. I. Genetics Definition Mendelian Genetics Drosophila melanogaster: The Fruit Fly Historical view of the fly Drosophila as a model organism II. Developmental Biology Definition Historial view III. Growth Control: The different parameters Our system: the fly wing Systemic vs Organ-autonomous Growth Control Size Control and Human Disease Summary

3. I. Genetics

4. Genetics Genetics deals with the molecular structure and function of genes, gene behavior in the context of a cell or organism, patterns of inheritance from parent to offspring, and gene distribution, variation and change in populations is a discipline of biology, is the science of genes, heredity, and variation in living organisms GENETICS + ORGANISM EXPERIENCES = FINAL OUTCOME

5. Mendelian and Classic Genetics Gregor Mendel (1822 - 1884) observed that organisms inherit traits by way of discrete units of inheritance, which are now called genes studied the nature of inheritance in plants traced the inheritance patterns of certain traits in plants and described them mathematically

6. studied the segregation of heritable traits in pea plants Pisum sativum Discrete Inheritance and Mendel’s Laws 29,000 pea plants Grow easily, develop pure-bred strains, and control their pollination

7. Discrete Inheritance and Mendel’s Laws

8. Dominant trait Alleles: is one of a number of alternative forms of the same gene

9. Discrete Inheritance and Mendel’s Laws

10. 3:1 ratio diploid species: each individual has two copies of each gene, one inherited from each parent organisms with two different alleles of a given gene are called heterozygous organisms with two copies of the same allele of a given gene are called homozygous

11. Discrete Inheritance and Mendel’s Laws (WW) Purple (Ww) Purple (ww) White heterozygoushomozygous

12. Discrete Inheritance and Mendel’s Laws (WW) Purple (Ww) Purple (ww) White Genotype (set of alleles) Phenotype (observable traits) heterozygoushomozygous one allele is called dominant other allele is called recessive W W

13. Discrete Inheritance and Mendel’s Laws

15. 3:1 ratio

16. Discrete Inheritance and Mendel’s Laws

17. The Law of Dominance: In a cross between contrasting homozygous individuals, only one form of the trait will appear in the F1 generation - this trait is the dominant trait 1

18. Discrete Inheritance and Mendel’s Laws The Law of Dominance: In a cross between contrasting homozygous individuals, only one form of the trait will appear in the F1 generation - this trait is the dominant trait 1 The Law of Segregation: when any individual produces gametes, the copies of a gene separate so that each gamete receives only one copy (allele) - a gamete will receive one allele or the other 2

19. The Law of Independent Assortment: alleles responsible for different traits are distributed to gametes (and thus the offspring) independently of each other Discrete Inheritance and Mendel’s Laws The Law of Dominance: In a cross between contrasting homozygous individuals, only one form of the trait will appear in the F1 generation - this trait is the dominant trait 1 The Law of Segregation: when any individual produces gametes, the copies of a gene separate so that each gamete receives only one copy (allele) - a gamete will receive one allele or the other 2 3

20. Drosophila melanogaster

21. Drosophila melanogaster: the fruit fly

23. Charles W. Woodworth (1865 - 1940) 1900 – First to breed Drosophila in the Lab Historical view of Drosophila

24. Thomas Hunt Morgan (1866 - 1945) 1933 – Nobel Prize in Physiology or Medicine for the role played by chromosomes in heredity 1900 – Started to work with Drosophila (study of mutation) 1910 – First mutation was found (white) Historical view of Drosophila 1911 – Genes are on chromosomes

25. Historical view of Drosophila

26. Hermann Joseph Müller (1890 - 1967) 1946 – Nobel Prize in Physiology or Medicine for the discovery of the genetics effects of Radiation (X-ray mutagenesis) Historical view of Drosophila

27. Eric Wieschaus (1947 - ) Janni Nusslein-Volhard (1942 - ) Edward B. Lewis (1918 - 2004) 1995 – Nobel Prize in Physiology or Medicine for revealing the genetic control of embryonic development Historical view of Drosophila

28. Jules A. Hoffmann (1941 - ) Bruce A. Beutler (1957 - ) Ralph M. Steinman (1943 – 2011) 2011 – Nobel Prize in Physiology or Medicine for the discovery of the dendritic cell and its role in adaptive immunity Historical view of Drosophila

29. Why Drosophila melanogaster is such a good model organism ?

30. Short Life Cycle (Temperature Dependent – 10 days @ 25ºC) Easy to maintain in the Lab (low cost) Suitable of Genetic Manipulation Simple karyotype: 4 pairs of chromosomes (3 autosomes + sexual chromosomes) Extensive set of genetic tools available Functional conservation of regulatory and biochemical pathways with humans Gene Sequence Conservation with humans: 60% Each Female lays 400-500 eggs

31. Why Drosophila melanogaster is such a good model organism ? Easy to maintain and manipulate in the Lab (low cost) Suitable of Genetic Manipulation Simple karyotype: 4 pairs of chromosomes (3 autosomes + sexual chromosomes) Extensive set of genetic tools available Functional conservation of regulatory and biochemical pathways with humans Gene Sequence Conservation with humans: 60% Short Life Cycle (Temperature Dependent – 10 days @ 25ºC) Each Female lays 400-500 eggs

32. Why Drosophila melanogaster is such a good model organism ? Easy to maintain and manipulate in the Lab (low cost) Suitable of Genetic Manipulation Simple karyotype: 4 pairs of chromosomes (3 autosomes + sexual chromosomes) Functional conservation of regulatory and biochemical pathways with humans Gene Sequence Conservation with humans: 60% Extensive set of genetic tools available Short Life Cycle (Temperature Dependent – 10 days @ 25ºC) Each Female lays 400-500 eggs

33. Why Drosophila melanogaster is such a good model organism ? Suitable of Genetic Manipulation Simple karyotype: 4 pairs of chromosomes (3 autosomes + sexual chromosomes) Functional conservation of regulatory and biochemical pathways with humans Gene Sequence Conservation with humans: 60% Extensive set of genetic tools available Easy to maintain and manipulate in the Lab (low cost) Short Life Cycle (Temperature Dependent – 10 days @ 25ºC) Each Female lays 400-500 eggs

34. Why Drosophila melanogaster is such a good model organism ? Suitable of Genetic Manipulation Simple karyotype: 4 pairs of large chromosomes (3 autosomes + sexual chromosomes) Functional conservation of regulatory and biochemical pathways with humans Gene Sequence Conservation with humans: 60% Extensive set of genetic tools available Easy to maintain and manipulate in the Lab (low cost) Short Life Cycle (Temperature Dependent – 10 days @ 25ºC) Each Female lays 400-500 eggs

35. Why Drosophila melanogaster is such a good model organism ? Suitable of Genetic Manipulation Simple karyotype: 4 pairs of large chromosomes (3 autosomes + sexual chromosomes) Functional conservation of regulatory and biochemical pathways with humans Gene Sequence Conservation with humans: 60% Extensive set of genetic tools available Easy to maintain and manipulate in the Lab (low cost) Short Life Cycle (Temperature Dependent – 10 days @ 25ºC) Each Female lays 400-500 eggs

36. Why Drosophila melanogaster is such a good model organism ?

37. Drosophila melanogaster Life Cycle Growth Phase

38. Drosophila melanogaster: why is such a potent genetic organism ? Mutant animals are readily obtainable Targeting gene expression in a temporal and spatial fashion Genome fully sequenced Huge amount of transgenic lines available

39. Driver lineResponder line Big collection of both Driver and Responder Lines available Temperature Dependence of the Driver Line Targeting gene expression: Gal4-UAS System

42. II. Developmental Biology

43. Developmental Biology

44. Historical Perspective – The first steps Aristotle (384 – 322 AC) Study of the Development of the chick The semen of the male provides the “form” or soul and the female the unorganized matter (menstrual blood) allowing the embryo to grow: EPIGENESIS Theory of Preformationism: organs with their own shape expand Theory of Spontaneous Generation: life of invertebrates emerges from non-living matter (“nothing”)

45. Views of a Fetus in the Womb Leonardo da Vinci, ca. 1510-1512 Dissection of human corpses Drawings of the vascular and system First drawing of the human fetus in the utero Historical Perspective - Renaissance Leonardo da Vinci (1452 - 1519)

46. Historical Perspective - Renaissance

47. Antonie van Leeuwenhoek (1632 - 1723) “…now that I have discovered that the animalcules also occur in the male seed of quadrupeds, birds and fishes…, I assume with even greater certainty than before that a human being originates not from an egg but from an animalcule that is found in the male semen” Discovered the microorganisms: animacules Discovered the spermatozoa

48. Nicolaas Hartsoeker in 1695 Historical Perspective - Renaissance PREFORMATIONISM organisms develop from miniature versions of themselves

49. Historical Perspective - Renaissance Discovered the follicles of the ovary (known as Graafian follicles), in which the individual egg cells are formed Reiner de Graaf (1641 - 1673) Rejecting the preformationism

50. Historical Perspective Ernst Haeckel (1834 - 1919) Recapitulation Theory / Embryological Parallelism developing from embryo to adult, animals go through stages resembling or representing successive stages in the evolution of their remote ancestors "ontogeny recapitulates phylogeny”

51. Opposing view that the early general forms diverged into four major groups of specialized forms without ever resembling the adult of another species Karl Ernst von Baer (1792 - 1876) Historical Perspective

52. August Weismann (1834 - 1914) Historical Perspective Germ plasm theory inheritance only takes place by means of the germ cells—the gametes Other cells of the body—somatic cells—do not function as agents of heredity

53. Historical Perspective Experimental Embryology Wilhelm Roux 1888 – Experiment destroying the frog embryo (in the two cells stage) Hans Driesch 1892 – Separates de early 4 cells stage embryo of the sea urchin Hans Spemann and Hilde Mangold 1918-1924 – Transplants of cells from one embryo to another induced particular tissues or organs – embryonic induction. Nobel Prize in 1935

54. Are Developmental Biology and Genetic Linked ?

55. III. Growth Control

56. How are differences in size achieved ?

57. What determines differences in size ? Size of an organ/animal = similar Size of an organ/animal = number of cells + size of the cells Cell Number Cell Size Cell Number + Cell Size Cell Division + Cell Death Cell Growth number of cells+ size of the cells+ space between cells

58. Cell Division / Proliferation: increase in cell number by one cell (the "mother cell") dividing to produce two "daughter cells" Cell Death / Apoptosis: is death of a cell in any form, mediated by an intracellular program (DNA fragmentation and protein degradation) Cell Growth: increase in cell mass (protein synthesis and organelle biogenesis) What determines differences in size ? Cell Cycle

59. How organs achieve a particular size and pattern ?

60. Drosophila imaginal discs: proliferative tissues

61. notum wing 20-30 cells 50,000 cells Drosophila wing imaginal disc Embryo Larvae Adult

62. Drosophila wing imaginal disc development

63. Body Size Regulation

64.  Cell autonomous growth promoters  Morphogens, signaling molecules  Long range signaling molecules (hormones…)  Environmental factors (nutrition…) Systemic vs organ-autonomous growth control

65. Systemic growth control SYSTEMIC GROWTH CONTROL GROWTH RATEDEVELOPMENTAL TIMING (moults+pupariation)

66. Gut Fat body Brain Ring gland nutrients Insulin GROWTH Systemic growth control FEEDING Hemolymph (fly ‘blood’) Ecdysone DEVELOPMENTAL TIMING

67. Organ-autonomous growth control Regeneration Experiments Transplants Experiments: when a small organ is transplanted into an adult organism it grows to its normal final size (even in between different species)

68. Size Control and Human Disease Cancer: tumor initiation, metastasis Diabetes and Obesity Organ hypertrophy or atrophy Insulin pathway dMyc oncogene Hippo pathway TGF  signaling (Dpp) Wnt signaling (Wg) Regeneration and Stem Cell Biology Growth Pathways Drosophila was, is and will be important for Human Biology

69. Crazy about B omedicine Thank you Development and Growth Control Lab

71. Transformation in flies