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노랑초파리의 난자형성과정에 따른 egg chamber의 미세구조적 변화와 RNA 및 단백질 합성에 관한 연구
- 노랑초파리의 난자형성과정에 따른 egg chamber의 미세구조적 변화와 RNA 및 단백질 합성에 관한 연구
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- 대학원 생물학과
- 이화여자대학교 대학원
- 노랑초파리（Drosophila melanogaster）의 난자형성과정 동안에 일어나는 egg chamber를 구성하는 세로의 구조적 변화와 RNA및 단백질합성 양상을 전자현미경적 관찰, 생화학적 방법 및 자기방사법을 통해 연구하여 다음과 같은 사실을 밝힐 수 있었다.
초파리의 난모세포질은 난황축적을 전후하여 크게 변한다. 난황형성이전에는 다양한 세포내소기관들이 나타나는 반면, 난황축적이 시작되는 시기부터는 다양한 크기의 난황립이 세포질을 채우게 된다. 여포세포의 외부형태 역시 난황형성을 전후하여 변화를 보이는데,stage 7이전의 균일한 형태로부터, stage 10에서 egg chamber부위에 따라 가장 뚜렷한 세포의 분화가 나타난다. 여포세포에 의해 형성되는 난황막은 일단 subunit인 vitelline body가 합성된 수 이들의 융합을 통해 형성되며, 이때 난모세포의 표층에는 물질 이입에 관여하는 pit, coated vesicle, tubule 등의 구조물들이 관찰된다. 또한 영양세포에 면한 난모세포막에 많은 미세융모와 endocytotic vesicle이 나타나는 것으로 보아, 영양세포로부터 난모세로포의 물질 이동은 ring canal 뿐만 아니라 막을 통해서도 이루어짐을 알 수 있다.
In vitro의 egg chamber에서 일어나는 RNA와 단백질의 합성능을 보면, RNA의 경우 난황축적 이후의 모든 단계에 걸쳐 비교적 낮은, 일정한 수준을 유지하는 한편, 단백질의 경우는 다른 시기에 비해 여포세포의 역할이 큰 stage 11과 12에서 상당히 높은 전구물질의 uptake와 incorporation양을 보인다.
각 단계의 egg chamber에 누적된 단백질의 약 10~20%가 단계에 따른 독특한 출현 양상을 나타내는 반면, 새로 합성되는 단백질의 경우, 약 20~35%가 단계특이성을 보인다.
Egg chamber를 형성하는 세포 중, 여포세포에서는 RNA및 단백질이 난자형성 전 과정에 걸쳐 합성되며, 특히 RNA는 배선시기부터 합성이 시작되고 있다. 영양세포의 경우, RNA합성은 stage 3부터 시작되며 세포의 역할이 가장 큰 stage 10에서 가장 왕성한 RNA합성을 보이는 반면, 단백질 합성은 난황형성 이전에 일시적으로 일어나고 있다. 난모세포에서는 stage 10까지는 RNA및 단백질의 합성을 관찰할 수 없으므로, 적어도 stage 10까지는 RNA또는 단백질 합성에 관여하지 않는 것으로 보인다.
; Changes in the ultrastructure and patterns of RNA and protein syntheses in the egg chamgers during oogenesis of Drosophila melanogaster were investigated by electronmicroscopy, liquid scintillation counting, two dimensional electrophoresis and autoradiography.
1. The ooplasmic constituents were found to be changed through out oogenesis. In previtellogenic stages, various cellular organelles were observed, but at stage 10 when the vitellogenesis occurs actively, the ooplasm was filled with yolk granules of various sizes as well as with an increased number of annules of various sizes as well as with an increased number of annulate lamellae. Large yolk granules seemed to be formed by fusion of small yolk granules. At stage 13 when the yolk formation was completed, different types of yolk granules appeared.
2. The follicle cells which were known to play a significant role in the formation of vitelline membrane and chorion were also differentiated during oogenesis in their external features. In previtellogenic stages, the follicle cells were nearly square at the side view whereas in stage 10 egg chamber the follicle cells increased in height with a number of elongated projections on the cell surface.
3. The vitelline membrane was formed from stage 8 through stage 11. The vitelline bodies were synthesized in follicle cells and these bodies were fused with each other to form a compact vitelline membrane. During the vitelline membrane formation, various structures which were appeared to be involved in transportation of materials, were observed in the cortical region of oocyte.
4. The observation that the ooplasmic membrane confronting the nurse cells have many microvilli and endocytotic vesicles strongly that the materials could be transported from nurse cells into oocyte through membranes as well as through ring canals.
5. The synthesis of RNAs in vitro at each developmental stage of postvitellogenesis occurred at a constant and low level.
6. The incorporation of 3H-leucine into newly synthesized proteins in the egg chambers of postvitellogenic stages was influenced by the amount of uptake of 3H-leucine. The rations of incorporation to uptake were high at stage 11 and stage 12 at which the vitelline membrane and chorion were formed by follicle cells.
7. Electrophoretic analyses showed that proteins were accumulated and synthesized stage-specifically in the egg chambers during oogenesis since approximately 10-20% of the silver stained proteins and 20-35% of newly synthesized proteins appeared to be stage-specific. The largest number of newly synthesized proteins was detected in stage 11 egg chambers.
8. Autoradiography was used to localize the newly synthesized RNA and proteins in egg chambers during oogenesis. The RNA synthesis in follicle cells was first detected in germarium Region 2 whereas the protein synthesis began at stage 1. These results indicated that the synthesis of RNA occurred earlier than the synthesis of protein in follicle cells. The RNA synthesis in the nurse cells began from stage 3 and became most active at stage 10. Most of the RNA which were synthesized in nurse cells seemed to be a rRNA since the silver grains were found mainly on the uncleoli of nurse cells. The nurse cells seemed to be involved in the synthesis of proteins only at previtellogenic stages.
9. The synthetic activities in RNA and protein in oocytes were not observer during oogenesis until stage 10. From stage 12 egg chambers the silver grains for protein were detected in the ooplasm except on the yolk granules, suggesting a possibility that non-yolk proteins may be synthesized in oocyte after stage 11.
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