Bài giảng Tế bào gốc - Bài 5: Ổ tế bào gốc - Trần Hồng Diễm

Nội dung text: Bài giảng Tế bào gốc - Bài 5: Ổ tế bào gốc - Trần Hồng Diễm

1. REPORTS that it remained a cystoblast, two lacZ+ stem One of the three somatic cell types, cap cells control IGS cell number (Fig. 3, F and G). cells were present at the tip (Fig. 2D). These cells, interacted with stem cells in a manner that In contrast, terminal filament and cap cells did REPORTSstem cells were connected by an elongated fu- suggested they play a role in niche function. not change in the absence of germ cells. Somat- some, indicating that they were recently divided Over the 36-day period, the number of cap cells ic cell divisions continued in their vicinity as in that it remained a cystoblast, two lacZ+ stem One of the three somatic sistercell types,cells in capearly interphasecells control (4); the IGS fusome cell numberand GSCs (Fig. remained 3, F and closely G). correlated at about germaria that form in the absence of germ cells cells were present at the tip (Fig. 2D). These cells, interacted with stem cells wasin a orientedmanner inthat an unusualIn contrast, manner, perpendic-terminal filament 2.5 cap and cells cap per cells GSC did (Fig. 3A). Moreover, (21). Despite their presence near the GSC stem cells were connected by an elongated fu- suggested they play a role in ularniche to function.the anteriorlposterior not change (dp) in axis the (10).absence GSCs of germ were cells. observed Somat- to always make special niche, these dividing somatic cells did not be- some, indicating that they were recently divided Over the 36-day period, the numberThese of observations cap cells suggestic cell a specific divisions model continued for contacts in their with vicinity cap cellsas in that characteristically come GSCs. sister cells in early interphase (4); the fusome and GSCs remained closely correlatedGSC replacement at about (Fig.germaria 2E). After that one form GSC inis thealign absence with of the germ 'dp cellsaxis of the ovariole. The Because the number of cap cells correlates was oriented in an unusual manner, perpendic- 2.5 cap cells per GSC (Fig. 3A).lost, itsMoreover, neighboring stem(21). cell Despite divides perpen-their presenceGSC's hsomenear the remains GSC adjacent to the GSCI closely with the number of GSCs, we inves- ular to the anteriorlposterior (dp) axis (10). GSCs were observed to alwaysdicular make to specialdp axis, causingniche, a thesedaughter dividing cell to somaticcap cell cells interface did not dwing.most be- of the cell cycle. tigated whether they might function by pref- These observations suggest a specific model for contacts with cap cells that characteristicallyoccupy the environment come recently GSCs. vacated by the In contrast, the behavior of inner sheath cells erentially sending a dpp signal. Suitable an- GSC replacement (Fig. 2E). After one GSC is align with the 'dp axis of thedeparted ovariole. GSC. The For this mechanismBecause to thework, number the andof cap terminal cells correlatesfilament cells did not correlate tibodies to Dpp are unavailable, so we used lost, its neighboring stem cell divides perpen- GSC's hsome remains adjacentenvironment to the GSCI at the site closelyof the lost with GSC the must number be closely of GSCs, with GSCs. we inves- As germaria aged, terminal whole-mount in situ hybridization to deter- dicular to dp axis, causing a daughter cell to cap cell interface dwing.most ofcapable the cell of cycle.programming tigated the incoming whether cell they to mightfilament function cells decreased by pref- in number from an mine which cells at the ovariole tip express occupy the environment recently vacated by the In contrast, the behavior of innerbecome sheath a GSC cells rather erentially than a cystoblast. sending Our a dppaverage signal. of Suitable9.2 (3 days) an- to 5.0 (36 days) (Fig. dpp mRNA. These experiments detected low departed GSC. For this mechanism to work, the and terminal filament cells didobservations not correlate indicate thattibodies it is capable to Dpp of doing are unavailable,3A) and changed so we from used a linear to a ball-like levels of dpp mRNA in both cap cells and environment at the site of the lost GSC must be closely with GSCs. As germariaso, aged, and hence terminal that GSCswhole-mount reside in a true in stemsitu hybridizationarrangement (Fig. to 3,deter- B to D) (19). Likewise, the inner sheathcells, as well as higher levels in capable of programming the incoming cell to filament cells decreased in numbercell niche. from an mine which cells at therelative ovariole number tip expressof inner germarium sheath prefollicle cells farther posterior in the ger- become a GSC rather than a cystoblast. Our average of 9.2 (3 days) to 5.0 (36The days) ability (Fig. of the dppovariole mRNA. tip to Theseact as aexperiments (IGS) cells detected and GSCs low varied (Fig. 3E). How- marium. No dpp mRNA was seen in terminal observations indicate that it is capable of doing 3A) and changed from a linearstem to cella ball-like niche is likelylevels to be biologicallyof dpp mRNA im- inever, both the capnumber cells of IGSand cells was closely cor- filament cells or in any germ line cells, in- so, and hence that GSCs reside in a true stem arrangement (Fig. 3, B to D) (19).portant. Likewise, Females the produce inner eggs sheathcells, for months, de- as wellrelated as higherwith the levelsnumber in of differentiating germ cluding GSCs (Fig. 4A). These results show cell niche. relative number of inner germariumspite the 4- sheath to 5-week half-lifeprefollicle of an cells individual farther cellsposterior (r = 0.88). in the A ger-functional connection be- that cap cells are one of several cell types The ability of the ovariole tip to act as a (IGS) cells and GSCs varied (Fig.stem cell3E). (11). How- To investigatemarium. whether No dppstem mRNAcell tween was seenIGS cells in terminal and germ cell cysts has been located near the GSCs that express dpp. stem cell niche is likely to be biologically im- ever, the number of IGS cells wasreplacement closely occurscor- normally,filament we measuredcells or thein anypreviously germ line suggested, cells, becausein- ovariole tips that Moreover, it does not appear to be the ab- portant. Females produce eggs for months, de- related with the number of differentiatingnumber of stem germ cells andcluding somatic GSCs niche cells(Fig. in 4A).develop These without results germ show cells lack IGS cells (I I). sence of contact with a dpp-expressing cell spite the 4- to 5-week half-life of an individual cells (r = 0.88). A functional agingconnection females (Fig.be- 3). thatDuring cap the cellsfirst 5 areweeks one of Toseveral investigate cell thetypes role of IGS cells in adult that causes the posterior stem cell daughter to stem cell (11). To investigate whether stem cell tween IGS cells and germ cellof cysts adult has life beenthe average located number near of GSCs the per GSCs geharia that weexpress studied dpp.females carrying a hs-barn differentiate as a cystoblast. replacement occurs normally, we measured the previously suggested, because ovariolegermarium tips declined that fromMoreover, about 2.5 toit 2.0does (Fig. not transgene,appear to whose be the stem ab- cells can be induced to Our studies suggest a working model for a number of stem cells and somatic niche cells in develop without germ cells lack3A), IGS significantly cells (I I). less sencethan the of 50% contact reduction with adifferentiate dpp-expressing (20). Over cell the course of several GSC niche (Fig. 4B). We propose that cap cells aging females (Fig. 3). During the first 5 weeks To investigate the role of IGSexpected cells inin adultthe absence that of causesreplacement the posterior(P < days stem after cell heat daughter shock, GSCsto were lost and all are critical to the formation, maintenance, and of adult life the average number of GSCs per geharia we studied females carrying0.01). Replacement a hs-barn stemdifferentiate cells must asfunction a cystoblast. germ line cysts completed development and left regulation of the GSC niche. Cap cells and efficiently because the rate of stem cell loss the germarium. Such germaria also lost all IGS terminal filament cells form a characteristic germarium declined from about 2.5 to 2.0 (Fig. transgene, whose stem cells can be induced to Our studies suggest a working model for a does not increase with age (11,12). Some of the cells, further indicating that developing germ structure with sufficient internal surface area to 3A), significantly less than the 50% reduction differentiate (20). Over the course of several GSC niche (Fig. 4B). We propose that cap cells ovarioles that did lose a stem cell started with expected in the absence of replacement (P < days after heat shock, GSCs were lost and all are critical to the formation, maintenance, and three GSCs, because the number of such ova- 0.01). Replacement stem cells must function germ line cysts completed development and left regulation of the GSC niche. Cap cells and rioles declined over the same period. efficiently because the rate of stem cell loss the germarium. Such germaria also lost all IGS terminal filament cells form a characteristic does not increase with age (11,12). Some of theDrosophila cells, further indicating ovary that developing germ structure with sufficient internal surface area to ovarioles that did lose a stem cell started with A CB three GSCs, because the number of such ova- GSC~- h rioles declined over the same period. & CPC' A CB germ Line stem cells (CSCs, red), differentiating germ cells (pink) IGs7 c' v GSC~- hterminal filament cells (TF, brown) * & cap cells (CPC, green) inner germarium sheath cells (ICS, orange) somatic follicle cells (FC, blue) B EF\ / germ line cyst (C) CPC' cystoblast (CB) GSC A )CB fusomes (yellow), fusome shape correlates with germ cell stage elongated stem cell fusome (EF) IGs7 c' v TF i * round stem cell fusome (RF) 4 RF B EF\ / ' 7 -& '"" Science. 2000 Oct 13;290(5490):328-30. GSC A )CB CPC Fig. 1. Germarium structure and stem cells. (A) TF i 4 Diagram of a Drosophila germarium in cross Fig. 2. A niche at the ovariolar tip can replace lost stem cells. (A) Generation of marked shn mutant section indicating germ Line stem cells (CSCs, GSC clones. All cells (ovals) express arm-lacZ marker (red), except shn mutant clones generated by RF ' -& '"" red), differentiating germ cells (pink), terminal FRT-mediated recombination as shown. (B to D). Cermaria with a recently lost shn GSC, analyzed CPC 7 filament cells (TF, brown), cap cells (CPC, 7 days after a heat shock to induce recombination, display two CSCs (numbered), indicating green), inner germarium sheath cells (ICS, or- replacement. For details of arm-lacZ marker (anti-IacZ, red), germ cell fusomes, and somatic cell Fig. 1. Germarium structure and stem cells. (A) ange), somatic follicle cells (FC, blue), and fu- membranes (anti-Hts, green), see (78). The Lost stem cell has differentiated into a young 16-cell Diagram of a Drosophila germarium in cross• KhiFig. GSC2. A nichephân at thechia, ovariolar TB tipsomes canch (yellow).replaceị em lostnFusomeằm stem c shapeạ cells.nh correlates (A)filament Generation with cyst oftậ marked n(B andcùng C, shn dotted mutant(TF) ovals), but is still a cystoblast (large dotted circle) in (D), indicating a recent section indicating germ Line stem cells (CSCs, GSC clones. All cells (ovals) expressgerm arm-lacZ cell stage. marker (B) Asymmetric (red), except location shn mutantof loss. clones A new generated cell (2) occupies by the position of the lost GSC and is still connected to the remaining red), differentiating germ cells (pink), terminal và FRT-mediatedTB mũ (CPC) recombination được asstemgi shown.ữ cell l ạ(Bandi to cystoblastđ D).ặc Cermaria tính relative withTBG, to asomatic recently TB ch lostwild-typeị shnem GSC, GSCvừ (1)analyzeda byr anời extended fusome. (E) An explanatory model for GSC replacement. A GSC filament cells (TF, brown), cap cells (CPC, khỏ7 idays TB after mũ ab heatắt đshockầu bitoệ cells.inducet hoáC, germ recombination, nh lineư cyst; là CB, các cystoblast;display cystoblast two EF, elon-CSCs (numbered),differentiates (CB) andindicating moves away from the cap cells (Left). The other GSC divides perpendicular to the green), inner germarium sheath cells (ICS, or- replacement. For details of arm-lacZgated stemmarker cell (anti-IacZ, fusome; RF, red), round germ stem cell cell fusomes, alp axis and (center). somatic Both cell daughters become GSCs, whereas the lost CSC is now a four-cell cyst (right). ange), somatic follicle cells (FC, blue), and fu-• Khimembranes GSC b ị(anti-Hts, mất haygreen), bi seefusome.ệ t(78). hoá The sLostẽ stemđượ cellc thayhas differentiated thế 1Bar intocách (B), a10 young pm.có 16-cellhiệu somes (yellow). Fusome shape correlates with cyst (B and C, dotted ovals), but is still a cystoblast (large dotted circle) in (D), indicating a recent germ cell stage. (B) Asymmetric location of quảloss. b ởA inew 1 cellGSC (2) bênoccupies cạ thenh. position of the lost GSC and is still connected to the remaining stem cell and cystoblast relative to somatic wild-type GSC (1) by an extended fusome. (E) An explanatory modelwww.sciencemag.org for GSC replacement. SCIENCE A GSC VOL 290 13 OCTOBER 2000 cells. C, germ line cyst; CB, cystoblast; EF, elon-• GSCdifferentiates bên cạ nhand movesnày phânaway from bào the t capạo cells nên (Left). TB The ch otherị em GSC đ dividesể lấp perpendicular đầy chỗ toTBG the gated stem cell fusome; RF, round stem cell bị alpmấ axist. (center). Both daughters become GSCs, whereas the lost CSC is now a four-cell cyst (right). fusome. Bar (B), 10 pm. • TB chị em trở thành GSC nhờ tín hiệu trong ổ hoạt hoá www.sciencemag.org SCIENCE VOL 290 13 OCTOBER 2000
2. REVIEWS maintenance. Finally, decapentaplegic (dpp), which stem-cell maintenance (see below), the absolute encodes a transforming growth factor-β (TGF-β)-like requirement of Yb for hh expression in cap cells fur- signalling molecule, is also essential for germline stem- ther indicates that cap cells might be the key signalling cell maintenance15.Although the soma dependence of cells. Third, ablation of most of the terminal filament dpp has not been tested by clonal analysis and its puta- cells increases the rate of germline stem-cell division 6. tive expression in cap cells awaits further confirmation, This indicates that only cap cells, possibly together dpp is expressed in inner sheath cells (FIG.1),other poste- with a few cells in the base of the terminal filament rior germarial cells and possibly in cap cells16. proper, might be required for germline stem-cell maintenance.So,cap cells probably constitute the cen- The central role of cap cells. In the stem-cell niche, cap tral component of the niche that is required for stem- cells seem to have a crucial function in regulating the cell maintenance in the germline. The fourth line of self-renewing ability of germline stem cells. Although evidence comes from the study of the DE-cadherin and this statement cannot be tested directly by ablation or arm genes, as mentioned above14.Although these pro- clonal analyses for the reasons mentioned above, it is teins are ubiquitously present in the germarium, their supported by four lines of evidence. First, cap cells are preferential accumulation at the cap-cell–stem-cell in direct contact with germline stem cells17;failure in interface and their requirement for germline stem-cell anchoring one pole of the stem-cell spindle towards maintenance also support the central role of cap cells the cap cells affects the asymmetry of the stem-cell in the niche. division18.Second,mutations in Yb completely elimi- As expected, the size of the germline stem-cell niche nate hh expression in cap cells, but only partially affect is correlated with the number of stem cells in the 9 GSC niche – con đường tín hiệu trong ổ hh expression in terminal filament cells .Because the germline. Germaria with one, two and three germline REVIEWS Yb-dependent expression of hh is essential for somatic stem cells have niches that contain an average of 4.2, 5.3 stem-cell maintenance and is involved in germline and 6.6 cap cells16.When a stem cell is lost,another stem Yb and Piwi seem to be part of a different sig- Cap cell (FIG.1) Yb nalling pathway .This is based mainly on the Box 2 | Germline and somaticovary stem cells and their niches in the Drosophila ovary following observations. First,Yb is required for the expression of Piwi and the Hh signal in cap and ter- The Drosophila ovary is composed of 9 ab minal filament cells ,but is not required for dpp Piwi 16–20 functionally equivalent expression (A. Szakmary and H.L., unpublished data). structures known as ovarioles (shown Hh This indicates that dpp expression might not be regu- in panel a).Each ovariole consists of Germarium Terminal filament lated by Yb. Second, although Yb and piwi mutants an anterior structure called the Arm have a germline stem-cell depletion phenotype that is GERMARIUM and a string of IS similar to dpp mutants,overexpression of Yb or piwi progressively more differentiated egg in somatic cells increases the number of germline chambers that reside posterior to the Cap cell Cap cell 9,13 ? stem-like cells by 2.5-fold .This increase,unlike the germarium.The tip of the germarium Dpp Hh contains a single stack of postmitotic Zpg tumorous phenotype caused by dpp overexpression, Egg ? Smo Hh somatic cells called terminal filament chambers III Ptc corresponds more proportionally to the increase in the Yb and piwi expression levels that are induced by cells (shown in panel b,a magnified Arm view of the boxed region of the IS IS heat-shock. So, Yb/piwi-mediated signalling from the germarium in a).At the base of the Nucleus niche seems to modulate the rate of germline stem- terminal filament cells are several Arm cell division in a dose-dependent fashion, whereas dpp squamous epithelial cells: these cap IS might operate through a different mechanism,possi- the underlying 2–3 germline stem cells GSC GSC bly by regulating the asymmetric fate of the two Self-renewing divisions Hh daughter cells. (GSCs),and are therefore called cap CB CT cells. Cap and terminal filament cells Smo What are the signalling molecules that are produced GSC Ptc form the stem-cell niche.A GSC Nucleus by the Yb/Piwi-mediated pathway? The Yb and Piwi divides asymmetrically with respect to proteins themselves are unlikely to be signalling mole- Arm its niche: one daughter cell will remain Nucleus cules.Yb is a novel protein that does not contain obvi- Ci SIGNAL PEPTIDE in contact with the niche as a stem cell, Oviduct Nucleus ous transmembrane domains or a .Piwi is while the other daughter cell will leave SSC SSC the founding member of the Piwi (also known as the niche and differentiate as a Argonaute) family of proteins, which encode basic pro- cystoblast (CB)17,18.This divisional Ci teins with homology to EIF2c (a eukaryotic transla- asymmetry is ensured through the tional initiation factor) and have crucial roles in stem- anchorage of one pole of the mitotic spindle by a cytoplasmic organelle called the spectrosome (the small orange spheres Nat Rev Genet. 2002 Dec;3(12):931-40. cell division, gametogenesis and RNA silencing in the 18,64 Self-renewing divisions 12,19–23 GERMARIUM in panel b) . SSC animal and plant kingdoms .A mouse homologue The apical, corn-shaped part of After GSC division, the cystoblast undergoes four rounds of mitosis with incomplete cytokinesis to generate a 16-cell of Piwi (Miwi) forms a complex with the mRNA of its the insect ovariole (the cyst (CT). Meanwhile, 2–3 somatic stem cells (SSCs), which reside in the middle region of the germarium, produce ‘pre- target genes to regulate spermiogenesis23.In the functional unit of the insect follicle cells’that form an epithelium surrounding the 16-cell cyst, to produce an egg chamber (not shown)35.SSCs are Figure 1 | Somatic signalling pathways in the Drosophila ovarian stem-cell niche. A model that summarizes the known niche–stem-cell interactions (see main text for details). Drosophila germarium, Piwi is a nucleoplasmic protein ovary) that contains both separated from their signalling source• by 2–5Dpp inner sheath signal cells (ISs), which might-> mean đthati theề nicheu forhoà SSCs extends số phận 2 TB chị em trong PBBĐX germline and follicle stem cells Innexin 4 (or Zero population growth, Zpg) exists at the interface between germ cells and also in both germline and somatic cells, including cap and and is responsible for the beyond the signalling cells alone (see main text).As the egg chamber buds off the germarium,some pre-follicle cells form between germ cells and follicle cells (not shown). Arm, Armadillo; Ci, Cubitus interruptus; terminal filament cells13.Piwi might therefore be continuous production of new an inter-follicular stalk that connects• the newlyYb formed/Pwi egg chamber-mediated to the germarium. So, the germlinepathway and somatic stem -> điềDpp,u Decapentaplegic;hoà tỉ GSC,lệ germline phân stem cell; Hh,bào Hedgehog; GSC IS, inner sheath cell; involved in RNA-related activities in the nucleus and so, egg chambers. cells together are responsible for the generation and maintenance of the ovary as an organ. Ptc, Patched; Smo, Smoothened; SSC, somatic stem cell; Yb, female sterile (1) Yb. like Yb, it is probably not secreted. • Yb/Pwi Hh-mediated pathway -> Ivai and II, type tròI and II receptors nh ofỏ Dpp, trong respectively. phân bào GSC A known signalling molecule produced by the Yb/Piwi-mediated pathway is Hh (FIG.1).Hh is specifi- | GENETICS | | 933 NATURE REVIEWS • Cell-cell adhesionVO LUME 3 DECEMBER mechanism: 2002 Zpg -> cần thiết cho sự sống cally expressed in niche cells10;a deficiency in hh causes © 2002 Nature Publishing Group cell will divide symmetrically along the cap cells, repop- germline stem-cell defects at a low frequency (20%), sót và biệt hoá TB mầm GĐ sớm ulating stem cells in the niche16.These interesting obser- whereas overexpressing hh in somatic cells causes a vations lend further support to the idea that cap cells are slight (56%) increase in germline stem-cell numbers9. the central component of the niche. This indicates that hh has a minor role in germline stem-cell division. hh expression in the niche cells Yb is required for both germline and somatic stem-cell divisionsSignalling from the niche. The molecular nature of requires Yb,but not Piwi9,indicating that Hh and Piwi somatic signalling from the niche is being revealed at a represent bifurcating branches of the pathway down- The piwi-mediated pathway is at least required primarily forrapid pacegermline (FIG.1).Dpp isstem-cell the Drosophila homologue lineage of stream of Yb (FIG.1).Interestingly,forced expression of bone morphogenetic protein (BMP) 2/4. Loss of dpp hh in a Yb mutant or overexpression of hh in a piwi he hh-mediated pathway is required primarily for somaticfunction stem-cell causes the divisiondepletion of germline stem cells, mutant can rescue the Yb and piwi mutant phenotypes, whereas heat-shock-induced dpp overexpression pro- respectively9.Therefore,Hh could be a safeguard mech- adherin-mediated cell adhe- sion is a general mechanismduces for germline anchoring tumours that stem contain numerouscells tocells anism that is downstream of Yb and can compensate for 15 SIGNAL PEPTIDE that resemble stem cells .Genetic clonal analyses on the Piwi deficiency.The germline stem-cell defect in Yb and their niches. (or signal sequence).A short, receptors and downstream transducers of the dpp sig- piwi mutants can also be rescued by generating patched 15–30-amino-acid sequence on nal,such as punt (which encodes a type II receptor) and (ptc)-deficient germline stem cells in Yb or piwi a newly translated polypeptide Mothers against dpp (Mad)/Medea (Med) (which mutants9.As Ptc is the receptor ofHh,and Ptc activity is that functions as a signal for its encodes a dimeric transcription activator), shows that repressed by Hh,the results of the rescue experiments secretion from the cell. The signal peptide is removed as the the Dpp signal is received directly by germline stem indicate that Hh exerts its function by acting through protein is secreted. cells. Ptc on the germline stem cells. 934 | DECEMBER 2002 | VO LUME 3 www.nature.com/reviews/genetics © 2002 Nature Publishing Group