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BIOLOGY 205/SECTION 7 DEVELOPMENT- LILJEGREN Lecture 2 MODEL ORGANISMS USED TO STUDY DEVELOPMENT 1. A brief introduction to six model systems widely used in labs today a. Why model organisms? Kit example b. Megabase genome =1,000,000 (million) base pairs; Gigabase = 1 billion! 2. Rules of evidence There are three main types of evidence that are accepted in all of experimental biology when figuring out a gene’s function. It is important to KNOW THEM: • Correlative Evidence o You see a gene expressed in a tissue o Weak, but relatively easy • Loss of Function Evidence o You knock out the gene, the tissue fails to form. o Better, but a little harder. • Gain of Function Evidence o You express a gene in an inappropriate tissue, and transform it. o Best, but hardest of all Or to sum it all up, think of the motto: “Find it, Lose it, Move it”. [but use the formal terms above in an exam situation!!] Oogenesis Behind every successful embryo stands a hard-working mother. 1. While both parents contribute DNA (the developmental blueprint), the mother also contributes two key ingredients to the egg a. a store of supplies providing for embryogenesis b. asymmetric cues to establish embryo's anterior-posterior & dorsal-ventral axes 2. Eggs are assembled in a complex, multi-stage process called OOGENESIS. a. Eggs are very large cells! They range from 100 µm (humans) to 1 mm (frogs, fish, fruit flies) to 1-10 cm (birds and reptiles). So even human eggs, although they seem small in comparison are 5x a typical somatic cell (20 µm) b. Eggs contain large stores of nutrients, called the yolk. Allow at least earliest embryogenesis & often entire process. c. Eggs contain large stores of macromolecules - machinery to synthesize & package DNA & assemble cells. (macromolecules include mitochondria, RNA/DNA polymerases, ribosomes) d. Eggs are enclosed in a coat or shell to: i. protect contents from damage & desiccation ii. regulate sperm entry. 3. In many animals eggs & sperm- the "germ cells"- arise from a special cell lineage known as the germ line. Gonads also contain specialized somatic cells. a. The germ line lineage often is set aside very early in development. Its descendants are the only cells able to generate a complete animal. Often specific determinants are segregated to specific embryonic cells which go on to form the germ line. This is an example of the segregation of determinants we talked about last time when we discussed how cells can asymmetrically receive info from their parents=P granules in C.elegans. [in figure, DNA stained blue using Hoescht dye, P-granules labeled with a fluorescent antibody to a particular protein found in them] Question - How does one single egg cell make all the stuff needed to start development? Answer - with support from her friends... Lets look at how eggs are assembled in flies b. In Drosophila, mitotic sisters (nurse cells) synthesize macromolecules and pump them into the egg. Germline stem cells divide 4 times, leading to 16 cells interconnected by cytoplasmic bridges. So the nurse cells are mitotic sisters of the oocyte. c. Nurse cells make macromolecules that are delivered to the developing oocyte through CYTOPLASMIC bridges (like gap junctions only larger!) i. Components of protein, RNA, & DNA synthetic apparatus ie. ribosomes ii. mRNAs differentially localized to particular parts of the oocyte. We’ll come back to how asymmetric distribution of certain RNAs helps to establish the anterior-posterior axis in the embryo in a minute. d. Somatic cells called follicle cells form an epithelium that surrounds egg. i. Produce nutrients that get pumped into the nurse cells and oocyte via GAP JUNCTIONS ii. Regulate import of other nutrients iii. May secrete eggshell iv. May regulate egg development by cell:cell signaling. e. All this help also makes it easy to have asymmetric (uneven) distribution of egg contents that can influence embryonic development. 4. Asymmetric distribution of egg contents influences embryonic development. Example #1- Fruit fly oogenesis. a. Molecules asymmetrically localized into one cell, the oocyte, before fertilization b. Nurse cells deliver mRNAs to the developing oocyte through CYTOPLASMIC bridges i. mRNAs differentially localized to particular parts of the oocyte- the front or anterior end (bicoid) and the back or posterior end (nanos). This asymmetric distribution helps to establish the anterior-posterior axis in the embryo. c. Localized maternal mRNAs also help establish body axis in vertebrates. Ie. Vg1 mRNA is asymmetrically localized d. Follicle cells also regulate egg development. Specific follicle cells exchange signals with regions of the underlying oocyte, to help establish the future dorsalventral axis of the embryo. e. For example, during development the nucleus of the oocyte travels to what will be the dorsal side of the embryo. Gurken mRNA is transcribed and translated into gurken protein by the oocyte but is localized only on the dorsal side. This signal interacts with torpedo protein (produced by the follicle cells), which causes these follicle cells to take on a dorsal cell fate, and inhibits pipe protein synthesis in these cells. On the ventral side, the follicle cells don’t receive Gurken (it doesn’t diffuse there), so they produce pipe, and a signal transduction pathway is activated to cause ventral cell fate. [There is also evidence that Gurken mRNA is also transported to the oocyte by the nurse cells, so this suggests that some dorsal/ventral signaling cues are present before fertilization and transcription of the Gurken oocyte genes.] 5. Physiological control of egg maturation- Example #2- Human oogenesis. a. Primary oocytes mature to prophase of meiosis I during embryonic development. They arrest at this stage, surrounded by a single layer of follicle cells, or primordial follicle. {2 million present at birth but only ~400 used during a woman’s lifetime} b. A fraction mature to developing follicles: the oocyte is enlarged & has a shell called the zona pellucida, & the follicle cells are multi-layered. c. After puberty, monthly surges of Follicle-stimulating hormone (FSH) from pituitary gland leads to growth of a group of developing follicles and the formation of receptors on follicle cells that recognize Luteinizing hormone (LH). Slightly later, the pituitary produces LH, which leads to follicle maturation d. LH production regulated by environmental cues (e.g. day length) in most mammals. Most mammals go through this cycle once/year, humans go through it monthly. e. Maturation is the completion of meiosis I & release of a single (usually) mature follicle from surface of the ovary, down the fallopian tube for potential fertilization. f. Follicle cells then produce estrogen & progesterone which cause Uterine & cervical changes, and feedback leads to a reduction in FSH. g. What hormone do pregnancy tests measure? hCG (human chorionic gonadotropin) h. RU486, a drug used to terminate pregnancy, blocks progesterone receptors and stops uterine wall from thickening and prevents implantation of the blastocyst. The hormone progesterone is needed to support pregnancy, so this drug can be effective up to 49 days. Interesting misconceptions about sperm: Anton van Leeuwenhoek, the famous Dutch microscopist who discovered sperm (1678) thought they were parasites living in the semen! Nicolas Hartsoeker, another Dutch microscopist and co-discover of sperm, thought each sperm contained a preformed human or “homunculus”. Leeuwenhoek believed that sperm were like seeds and that females merely provided ‘the soil’. 6. SPERM are small and motile and made by the male. a. Unlike eggs, Sperm do not need all the nutrients and building blocks, they need to MOVE to get to the large sluggish eggs. So they develop differently. b. SPERMATOGENESIS differs from oogenesis in several ways: i. Cells enter meiosis CONTINUALLY from puberty on. ii. Each cell that starts meiosis gives rise to 4 GAMETES, not just one iii. Much of sperm differentiation occurs after meiosis is complete. iv. Sperm differentiation makes a highly specialized structure with a head and tail v. it can move vi. in the head is an acrosomal vesicle important for accessing the egg NEXT LECTURE: sperm and egg hook up