LIFE CYCLE OF DICTYOSTELIUM DISCOIDEUM

LIFE CYCLE OF DICTYOSTELIUM DISCOIDEUM

l 1. HISTORICAL Cellular slime molds were first discovered by Brefeld in 1869 and the uniqueness of their asexual life cycle was first recognized by the french mycologist Ph. van Tieghem in 1880 (Bonner, 1982). Dictyostelium discoideum, a new species of cellular slime mold was discovered by Raper 1n 1935. The discovery of this organism which was especially suitable for experimental work gave a boost to research on cellular slime molds. The wild-type strain (NC-4) was unable to grow axenically and they required bacteria as food source. With the development of axenic strains (Ax2 and Ax3) which could grow in defined medium (Sussman and Sussman, 1967). Dictyostelium discoideum has emerged as a.major tool in the study of various biological processes. 2 CLASSIFICATION There is considerable disagreement among biologists regarding the classification of cellular slime molds. Some prefer to consider them as fungi while others place with protozoa. Raper {1982) and Grell (1972) classified cellular slime molds as follows. K.B.Raper (from Bonner, 1982) K.G. Grell (from Grell, 1972) Kingdom Plantae Kingdom Animalia Sub Kingdom Mycetae Phylum Protozoa 23

Division Myxomycota Class Rhizopoda Class Acrasiomycetes Order Amoebina Sub class Dictyostelidae Sub order Acrasina 3. OCCURENCE OF DICTYOSTELIUM DISCOIDEUM Cellular slime molds are found all over the world from the colder temperate regions to the tropicals. Dictyostelids are most abundant in forest habitats that are not too moist. They are rare or absent in dry, desert habitats (Raper, 1982). The amoebae of Dictyostelium discoideum inhabit soil, humus, and animal dung, and normally feed on bacteria. 4. LIFE CYCLE Dictyostelium discoideum has two alternate life cycles: sexual and asexual. The asexual cycle is the one which is used for almost all experimental studies. The life cycle involves a growth phase consisting of vegetative amoebae and development phase 4.1 Vegetative phase During the grqwth phase cells exist as individual amoebae. The amoebae are haploid with 7 chromosomes. These cells respond to factors such as folic acid which is released from bacteria, their normal food source (Bonner, 1982). The amoebae can be grown on agar plates with bacteria as food source or in axenic medium 24

l 4.2 Development phase When food becomes scarce, some of the cells start to secrete a chemoattractant, and cellular agglomerates are formed by means of chemotaxis (Bonner, 1982). During the process of development a multicellular fruiting body is formed. This process involves various stages like streaming, aggregate formation, and slugs. The slugs differentiate in to fruiting bodies. 4.2.1 Cell streaming When deprived of nutrients Dictyostelium discoideum cells cease growth and start aggregating. The aggregation of the cells is organized by the extracellular adenosine 3, 5 cyclic monophosphate (camp), which mediates chemotaxis and oscillatory cell-cell signaling (Devreotes, 1989). During development the aggregation competent cells produces camp. This camp is sensed by the distal cells and they in turn start moving towards the source camp. When camp binds to the distal cells it activates adenylate cyclase. These cells now releases camp which causes activation of more and more cells. This relay of camp causes the cells to stream towards the aggregation center. During the process of adaptation the adenylate cyclase is turned off. It regulates the level of camp. In Dictyostelium discoideum there are three types of phosphodiesterases, ( 1) membrane bound, (2) extracellular and (3) intracellular.. During 25

deadaptation the exc.ess camp is hydrolyzed by extracellular and membrane bound phosphodiesterase {Gerisch, 1987). 4.2.2 Cell-adhesion Chemotaxis and signal cellular agglomerates. Multi relay leads to the formation. of cellularity is maintained by the expression of two types of cell-cell adhesion sites. These two types of intercellular adhesion sites differ in their sensitivity to EDTA {Siu, 1990). The EDTA-sensitive sites were named contact sites B. They appear early in the developmental cycle and are easily disrupted in the presence of 1 to 2 rnm EDTA. The EDTA - resistant sites were named contact sites A. They are stable in the presence of EDTA up to a concentration of 15 mm and they are expressed by cells at the aggregation stage of development {Gerisch, 1980). 4.2.3 Cell-cell adhesion molecules in Dictyostelium discoideum Several surface glycoproteins have been identified as cellcell adhesion molecules at different stages of Dictyostelium development. a)glycoproteins (1)gp 24 The EDTA sensitive cell-cell binding is mediated by a cell surface glycoprotein of Mol. wt 24 KD. It is synthesized maximally during the first 4 hours of development (Siu, 1990). 26

(2)gp 80 gp 80 is involved in the EDTA stable cell-cell adhesion (Siu, 1990). gp 80 is synthesized mainly during the aggregation stage. (3)gp 95 The synthesis of gp 95 begins at the aggregation stage and continues throughout development. It may take over the function of gp 80 during the later stages of development (Siu, 1990). (4)gp 150 gp 150 accumulates between 6 and 18 hours of development. It has been suggested that gp 150 may play a role in the sorting out of the prespore and prestalk cells during pattern formation (Siu, 1990). (b) Discoidins Discoidins are galactose binding proteins suggested to involve in cell cohesion. The two types of discoidins, discoidin 1 and discoidin 2 are produced at similar times of development (Alexander, et al, 1983). 4.2.4 Slug stage camp chemotaxis results in the formation of an aggregate. The center of the aggregate is at first rounded but later forms a small tip and rises in to the air as an elongate cylinder. Under favorable conditions, the cylinder of cells assume a horizontal orientation and migrate. The migrating slug or pseudoplasmodium has an anterior half containing prespore cells (Bonner,1982). 27

4.2.5 Culmination and fruiting body formation At the beginning of culmination the slu~tip stops forward movement and points upward while the posterior end continues to move so that all the cells are gathered directly under the tip. At this stage, the stalk cells begin to form near the tip at the '\.) upper end of the prestalk zone. The stalk cells move in to the open upper end of the stalk by a reverse fountain movement. This causes the apical stalk initial to be pushed down through the prespore zone. The whole cell mass lowers and bulges laterally forming the mexican hat stage. By the continued elongation of the stalk the prespore mass rise in to the air. The fruiting body thus formed is called a 'sorocarp'. The pres pore cells differentiate in to spores. The spores are of 5 to 7 micro:::::.meter size and are liberated by the bursting of the sorus. The spores on germination produces the vegetative amoebae. The life cycle of Dictyostelium discoideum has been depicted in Fig: 1. 28

FRUITING BODIES 2 5 X ( 2 4 h rs) t SPORES GERMINATION t 200 X )... \ \ - ( SLUG 5 25X(18hrs) t ') AMOEBAE 320X STARVATION ' AGGREGATES 50 X ( 12 hr s). ", CELL STREAMING SOX!Bhrsl Fig. 1 LIFE CYCLE OF DICTYOSTELIUM DISCOIDEUM hrs - hours in development