$P ;5) & Lecture 1. Cornell University. *Tuesday, ijovember 16, 1965 h Aspects of Gene bontrol in Sigher Cr+nisms. I. Influence of bacterial and pha$;e genetic and molecular studies on genetic and biological conce_nts. 1. Basic genetic manterial: DW: its bases, its organization; its coding. 2. kode of operation of tile genetic system: DNA Heplic:ition- ~NH polymerase DNA transcription; KNii polymerase, base sequence of mRNA, mRiU transcriptions through ribosomes; transfer RNA, a?ssocisted enzymes. Proteins formed; specificities, 4. Types of genetic coi:lponents in the bacterial X& DNA: Structural genes Genes Genes Genes for transfer RNs for structural proteins for ribosomes: B. subtilis: fl f:Ij?! 2s - - 5. Regulation of acti:jn of genes: 1nduce.s and Repressors. The Cperator-- reading fr:,me of structural gene Regulator genes. Organization of structural genes: Cperons One Regultitor: Positions. Number og _:enes: thair relations Cperon plus others: one regulator. Arginine, Regulators: adjacent to operator: Lac locus. The activator component: Regulator: positive: Arabinose ABDC + E The C: Activator. The feedback mechanisms. 6. Regulation at the g~wa&i gene level: not yet clear how this operates0 7. The forgotten type of regulation: Salmonella -- will return to this la&r. the H and H2 duplicate genes in dhe host range: 1 fliKq&iw lviodification gf phage genome by host. f 8. Effects of episomes: *sol,:-ied episomes; the co:.trolling episomes (Austin) (Dawson). g . JNi?'O~~ik~~~ T : enormourstimultis The ac;ept$nce of differential r.:gulation of gene action: to reconsider mechanisms res:>onsible for control of gene action during diffe,*entiation of higher organisms. Although the basic components are the same: i)NA etc, the mechanisms controlling the action of the genes in higher org;;nisms are far more complic.%ted. This related to the very highly organized, co:+'iex bodies, the chroi::osor.es, and the highely organized nuclei, 'ihe extraordinary co-m lex organizatioil of the Chrol.iosoi,!es and nuclei geve?opment, rob bly related'%?'mechanisms controliing the ai:tiun of genes during - 2 - The various comr)onents of the chrti;:os..J-i.:es and the organized nuclei probably represent the components behaving like those of a computer: higi,ly organized set of wnsequ tive events, event: each related to the previous J = ? Example of this:@One must consider the caterpillar and 't ;tioth are extraordinary corqjjiex individuals but utilize the same the moth: set of genes. The computer: &/One must consider polymorphism: mimicry patterns, etc, the "Swithh Genes": One or Two mendelizing units switch the "compute$ from one sequence of interplay of events to another sequence, using the same complement of genes for this. -I ,dL*;i? #k ,~a.@ mq ,i& . 10. r`ie must consider that regulation of gene action in a nigher organism is a higb.iy programmed sequence of events from egg to mature individual. The large num-ber of different components of the chrcl.,osome are probably the component elexlents in this programed sequence. Question: Wh~L-t do we know about these components of the chromosome and what do we know about individual mechanisms in the system? II. The Chromosomes and their component parts: 1. Compared to bacterial chromosome, the breadth of the chromosome is enormous: clearly visible in the light mi:>rosoope. 2. Number 08 DNA molecules w,thin an individual chromosome: Very recent investigations, taking DNA from the chroeioso es, suggest that each chr:jr;osome has rel.ltively few DNA molecules. (8 to lo?) ( ;AJ 3. The non-DNA components of the chrmoso..:es: a>. The Histons: Ten different types known. Fall into 4 general classes: Ia, Ib, II, III, IX. 13ased an lysine-arginine ratic 8:l 1O:l 1.7:1 0.7:1 0.7:1 (No tryptophane) 1 - v@ y% ,, Same qualitative types of histones in all nucieated orgnisms. II II II II in active and in inactive chromoso::;es with very few exceptions. +q Some turn-over of hAones in nuclei that do not repliclte DNA Quantitative differences in hLstones but not yuaiitative differences. Considerable amitutiht of evidence that histones related to repression of gene action. %t, what substAnces a-:-e related to activation of gene? b). RN&4, new species, related to histoxLes. One of these RluA molecules for several of the histox:es. wufit /-3-?ti d. The residual protein - acid type; Phosopho proteins. RNA, new species, associ:ited with this protein, pwJ)'& , ,; ',?: i d). l?hosopholipids, - 3 - 4. The chro,:oso.;ile: an extraordinarly co!,lplex system: Related to control of action of genes; to mechanisms producing ~RNK etc.&&@, mechanisms of reduglicution of the dhromosomes, etc. We should expect to find that new, previously unsuspected mechanisms of protein formation, and repliczti,ns may occur with these strUctureso 5. Bacterial systems: do not have all of these. A few exceptions indicated and these may help in explaining some of the modes of regulation of gene action in higher organisms. The one outsttindin;,; example is the control mechanisms operating at the Ii1 &almoneila, and H2 duplicate genes in III. The relation of the organizatin of the Chr:Jwosoi!ie parts in the nucleus(to gene action and repression,,) GXKt$RsL Cu~~3I~LW~Iu~S. 1. The active chromatin: dispersed; the inactive chromatin contracted0 aj. (This realized by cytologists for many years: 33x:7mpie ia the l!Ecrospore of plants: b). Condensed nuclei and chr:imatin in generative cell; size of nucieolus. 2. The nucleglus organizer: Special region in one ChrG!.iosoi:;e. Function: to produce the nucleolus at telophase; The nucleolus with the DNA of organizer: related to production of the cytoplasmic ribosomes. l&w a-l~,~~~ Jxam@ie of the organizer: its position with respect, to the nucleus; the Constance of this osition in the working nucie;ls. Slide 1: Maize set of chrol,!oso es at pachytene. 6lide To -* The nucleolus chromosome in maize, 3. The true heterochromatic regions within a chromosoine. a>. The heterochronatin about centromeres in Drosovhila. The chromocenter in the working nucleus. Its position with-regerence to the nu:;lear membrane. ?? ??*?o????? ?O? o ? b). The true heterochromatin in maize: the knobs: Lliueb LO 4. The relation of the knobs to the nuclear membrane: Side 4 d). The relation of the centrome:?es to the nuciear membrane: bjiide 5 - ej. 3'he Betzrochromatin at the ends of chro:.os:j;;es: Very visible in many plants and animlas: delation to nu iear membrane. 4. None of these types of "heterocnromatin" h.tSe conventi.nal genes in them, `iy 5. The relation of cllr,:mat n carts to the nuciectr membrane is llighly significant as wi.11 be indic.%ced. -4- IV. Condensation of cnroi:iatin/\; one form of control of action of genes. Two distinctiy different classes or? this; e:-tch , however, is effectilae in repressing the action of structural genes, CLriSS I: 1. Two nucleated microspore: The active nucleus and the inactive nucleus0 org,nieation of chromatin in each. 3 -. Types of condensation of chro?.iatin in different nucLei of the same tissue: rabbit retinal cells in embryo; deermann photo. Slides 6, 'Jo -- 3. Calf thymus nucleus: Prenster, HllSiyey, I;lirsky Lab. 4. Appearance of nuaieus in light microscope before and after swelling, Slide 8. - Position of condensed chrGmatin in cor:Ldensed regi,ans Slide 9. - b j . 'Tests of parts of chromatin in such nuclei that produce mKU: Slide10 = d. The relj~tion of strands of DNA in association wit:; condensed regions: -@'igure of Prenster: Draw. d). Components of chromatin in active and inactive chronatin: Table 1, r'renster Play, 1965: 4. dffect of additions of polyanions to active and in:lctive chrmatin: Suggests that the special species of DJA ass ci%ted with the residualxMti protein associated with activation of gene whereas HMA s_uecies associ,ted with histones may reg:Jate in some manner repression of DKAo 4. The contracted chromatin: Specific types fbr elrch type of cell. 5 .QUESTIU~W: (1) bdhat type of mzchanisns controls these highly sl;ecific types of contractions of chromatin? (2) 'Why are the condensed parts bulked as they are with the active paL-ts extending from them in all parts of the c>..Gricted mass? (3) xhat is t- he relation of the nuc1e.r membrane to 311 of this? How does it come into the picture of repression? parts (4) What types of det-rminants coiltrol the particular of a chro:!oso!.le that will be i:: in any one type of cell? the cor.densed stti;re at any one ti-ii;e or (5) d&t combone:?ts of the advance so that they will becb:;le co:!tr--leted mechanism serves to rele:$se these settings? Are they particular elements associ-ited with the DNA-- the genes? and if so, what are these components? 03 If (5) is in fact correct, do we have any evidence for such settings and erasings of a setting? These questions indicate the extent of our ignoraace of the casitrol mechanisms in nuclL ated organisms ant the molecular level. de a,-e not yet ready to consider in detail any control mechanism in higher org;znisms at tnis level. !Ire must know more about the specifics of chrocosome co,iposition, organization of parts, and chan;es in this that occur0 CLASS II CUI\I.3~:1;1Y~i'i'I~~:as. This involves the compietB contqction of an entire chromosome or of a continuous segment of an entire chromosome in contrast to Class I which involves intermittent condensations within a chromosome, 1. Our knowledge of the control mechanisms better with this class. 2. The X chromosome in mammals as an example of this: (a). XY : X chroi;losome is not contracted. (b). X X: One X contracted; other X not contracted, (c). Position of the contr:tcted X in the nucleus: At the nucleiir membrane. Slide 11 (dj. Contraction of X when more than 6x0 present: xx, XXX, Lsxx, x.xiim, (e). Selection of which ii to contract: iTX female: Occurs during development: one X in one cell, &hd other X i. another cell, 'bVil1 co;sider this shortly after several other cases reviewed0 (gj. The CO&TRULLING ~L~-PM~N'T responsible for contraction allong chrol..osome 0 (1). Translocyitions bet;ueen ii and autosomes in the mouse: d region of co trol in the X chromosome. _ -. :I . the Hussel. Contraction occurs to either side of it, t5P~~*~#/~-J Distance controlled by this region Transloc :.tiors: autosome genes in line with this, they become contrticted and non-active: a c &&I ;g:T /\eN? : !I". Diagram: -.-.. :itricted. (b) Sperms: Set from father disc.dred during meiosis; only set from mother is carried in the s!:erm. (c). The female: regular meiosis: dome females: produce eggs that develop into females,, Under some cxditions, produce some eggs that develop into males. (a. 'ihe female embryos: both sets of chromos~~mes are functional and euchromatic. (e) The male embryos: One set--thet received i.rorn father, carried in the sperm-- becomes totally conden edf: Slide 12. Contracted chrornoso;ries again up against the nucleir membrane. Setting to do this occurred in germ line of male: then euchBomatic, (f) An.)ther group of coccids: same general co:lditions up to time of condensrttion of male set in the male embryos. Instead of condensation of set, all chromoso~les are elsminaued from the nuclei at a certain cleavage division. %* (g): conclude: some relationship between condensation and elimination process as these are related in evolution. ";yw-; L- 2 i;&& &-&I%+ ;$I!4 .WllfL4l. 4. The X for elimination chrom,s)i,ies in the cecidomyidae: (aj. tigg after ferti1izati.n: (b). fhe pole plasm: (c). Nucleus associ ted with pole plasm(germ line). Rescued from elimination, Any nucleus placed here: no eliminatiLn will occur. Elimination will occur if nucleus norma ly here rePlaced by one that would otherwise form soma and have chromoso,:es eliminated, in all nuclei (d).fChromosomesAsetfDr elimination at particular division in cleavage0 Kesude from this setting by contact of nucleus with particular ~toplasrnic component0 (Must keep the setting in advance for eliminatin and rescue of this by cytoplasmic component in mind for later use with ii chro!:iosomes in mammals: a rescue process,) 5. yo get all of this in focus, will consider the caxe of Sciara: controls of the behavior of the X chronosome: (a). 'The germ line of the male: 4 chrs Me stage in spermatogenesi6, from mo$$er, 4 from father at (b). Apermltogenesis: ii,l$iotic divisions: - 7 - (c). The two types of females: All with normal meiosis X'X = produces eggs, all of wrlich develop into females, xx = CW produce eggs, all of w-!lich develop into males, (d). The zygotes and early cleavage nuclei: Eggs from X's individuals: Cleavage: Zygote: X A mother/ 1K X A father Cleavage: 6 or 7th division, one X from father eliminated, JWbryo develops into a female, Eggs from & & mothers: Xygote: x A/ x x A: Cleavage: both X Cbromosoikes from male eliminated: develops into a male, I vlui (e) Transloc&tions between autosomes and X. Vlzrous ones worked with. Result: The X chromoso!ile: ~11 the aberrant events controlled by BLX~&H~~XH some element cttrried in the tiny short arm of X w -ch is heterochromatin, If transloc-tion occurs in short arm between centromere and Weterochratin, the chromosoL;e carr2;ing this and allFtof the autosome part: undergoes same events as X chromoso;,ie. *.*... r.... 1, - R P \ia.&~~:.& & ,+%a. .+ The heterochrom;itin carries a controlling element that accomplishes all of the events. How does it do this? (f). Transloc tions with aberrant disjuncti,ins:in the females: *'emale croducing females: 8/xt mothers: lew dggs : No li; oniy autosomes. Cons$it2tion of zygote: OA ii/ ii r: ii JQimination: One chro!>losome X only 1 L$~~~&LMJI ptid$ "~-4~~~.,~~ d i4 q&t I ;&.z8 Translocr~hti ,n from Xiit I mothers: i\i`ormaily male producers 1 \ some eggs: chror;osor:ies with controlling element of & i Constitution of zygote: tilimination: two X chron;;)soi.jes in cleavage(from father), Lxceptional females: retain two g chro:Ll js~, e, as one particle - Suggests: The X (not the X1) produces somes su.bs-cance.in eg:g before meiosis that can rescude one X from eliminc:.ti~jn Jith 1cir: females; two A chromosoI.es may be suck particles. Th.e:aefore 2 A rescued by it, - 8 - 6. The rel.Altion of Sci,za evidence to ii: chro:t!oso;l!e cunaens ixj-'tns in mammals. a>. ~11 X chrjmos ,:,. [es a:'e set in adv:ince to become cil,~densed duri g development of embryo. b). Gan be rescued from this by reYcti-!>n with some comborient in the cytoplasm. d. Clnly one such com:?onent produced. dj. This element reverses the setting for condens.itisn. 4. Should be matter of chance which X, if more than one present, will be rescued during stage when rescue occurs. f). XY - the ;C always r scued as no other h prese:it. xx - Une ir rescued in some ceils, other condenses; the other is rescued. In other cells, xxx, XLLi, nnnK: - oniy one 1x: can be rescued; all others are condensed. 7. Return to Sci.ra or the cocci.&: the germ line of the father. Setting of elements must occur in the germ lkne of the mother,, Erasing of the setting must occur in Xlements 8. Certain chroliosomal components .a:iae res;lsnsilale for the coiltrol of of contraction and reier+se from contriction; release from repression. for repression and These ele:nents respond to cytoplasmic or intranuciear substrnces, previously produced or introduced: as with hormones that react with the chromosomes themselves-- special p,irts. 9. If we accept that there ar4 chromosomal elements, distinct from the genes, that control their action $H% through v.rioGIs types of res onses of the chrjmatin m_te$ials, we are a long way on the road to unde.r*standlng the mechanisms thst operate in higher orgxnisms to control the action of genes. V. Other evidence of the manner by which the action of genes are cxltrolled: 1. Lactic dehydrogenase genes: H and M. Tetrameres: HhHH, HHHM, i%-INM, HMMN, NLXFi, Young and older individuals. (Ghick; rabl;it) 2. Intra-allelic repressian: Tetrahymena and karamoeciuq - J J - 3. Ijiuitip,.e genes - rJerotypes in r'aramoecium: tini:: 0:ie active at a tii&:e; otjlers inactive, . .I i rl I I. / 4. Aster;Ise alleles in maize: ':o:trol of actL!n of one or the other allele: Differences in different tissues, -+p Control mechanism .associ.zted with so:::et:lIng at Setting of a gene locus at one specific time in one tyi,e of cell to be expressed in cells some cell gerier:zt.I ,ns l`iter: Position effect: Becker w" iocus. Adjacent to he t zr0Cz clTU;it:.t in by trznsioc :ti.nn. Gene set in s ,yle cells at th:i.s time. 2lffec-t of extra Y: No effect on the time of setting. X'fects frqu! ncy of czls that hqve their locus set at this time, iieflects type of setting mechansims that occur during t?evelOiJmento T-6 6. Alleles th it control the pattern of distribution in a tisajue of the end produc:t of Sequence of f'eIle act `.V"s : Pigment 3s eiiiI:l-(`1 e. Lady Beetle: Pigment pat-terns in the ~lytra. Types of pattijrns obCjerved in nanntite. Genetic analysis: tine locus as:rociated with cor:trul of tnis ~:'ittirm. il;ach allele of this iocus res.,i;,lisib.!e for one prticular pAttern: for control of production of end prOdJet of genes in sequence to :j~l.gment formation. elemc:nt combination of two a;-ieles of t ,.s ~~re~ul~tor",a~w~x Slide 13 Werlar,:jing p-ltterns. IiX:~,dh expresses itself inde endel1tA.y oY tr:e-0th pro 'Thus , CytO~jl2SLl in t is case is not a de-terminL9g fiictor directly, Til;le of ;jej;e action of an ?.?lele, number 0-r' zeiis i.n ?J ch acti;.?n wi.1.L ooour controlled by some COYI ,*-jnent at t.1~~ ,;er:e locus. 7. baciy Beetle patterns: Can du:;;lic : -:e com.~:.e7;ely with ncii ze wh.:re $9 e know the element at the gene locus and how Lt operates to acc~m lish t'hese Iatterns. 'This will be co:lsidered in next lect.ueo VI. Conclusio~,s: There are p::.r'ticul~ar chr-,i.,osoi:ial i:OI!;!;Ofie!itS , cnr ~;,.USOiri;.il eiements, that are res, onaible for the c ,I-itroi of ge:le action and they ;-icc~r~!~~i~sh th:s in ~a.7 ous minners: contr -:stL,ns ana re_:-;-k,2e from co? tr::ct-;.ons; re-Fression ioc-! 1 iy itrld rel. e:li :- e from t?Ls reoresslon. The Urg di:lJfe.:*ent sites in the genome: Bacteria: the episomes. f>.j d Xigher organisms: Unly one cr*se of "transposition" of some comfonent and this one that contrlls sex0 r "~~id'j fct&&& A`\ &,,g&u33 Sex control mechanism in Piegaselia - a fly: lo 2. 3. 4. 5. 6. 7. 8. 9. slides. Lecture 1 dhromosome set at pachytene: maize Nucleolus chromosome at pachytene:maize Nucleolus chromosome, knobs, pachytene, maize Knobs at nuclear membrane; endosperm of maize, polyploid. P,I!eulgen stain, p:-ichytene maize; B-type chro-r;oso.:e centromere. Kitten, 10 day old retina cells: .ijeerman Same Lymphocyte nuclei: non-swollen; swollen II II swollen 10. II Active and inactive chromatin: autoradisgraph tes:,s. 11. Sex chromosome in embryo tissue of rabbit. 12. Gondensed set of chromosomes in spermstogenesis i;`erococcus (me.aly bug). 13. blytra patterns: m~exEk Lady Beetle, Tan, 196 I Cornell University, becture 1. Igesday, PJOV.?~, 1965 I, The influence of bacterial and phsge genetsc and molecular studies on genetic and biological concepts, 1. DNA - mRNA (r@ipolymerase) - transfer BNfi, ri$osoii,es etc, Ditt)$UU& 2. Types of genes: structural, sRNA, rRlJ;I, regulators, suppressors, Operator- operon. Regulation. 3. Regulatory mechanisms: not yet clarified, 4. The odd types of control in bacteria: Hl-H2 dup&icste genes: Phase wriation tipisome control of gene action: Taylor; Dawson. Host modification of phage:. Modification of phage genome - restriction of host. 5. Impor.ance of the "odd" types: particularly the Hl-fi2 dupiic,.te genes, II. Types of gene control that have been e.amined: Xxamples. 1. at the molecular level: iAaemoblobin in young and older individuals, Lactic dehydrogenase: Two genes, N,H, lhe tetrameres. Young vers older embryos. Xsterases in maize: Timing of action of different alleles: COiiStXlCy for an allele: Control: genetic methods, resides at the locus of the gene.h filpidnadyq~g. 2. raramoecium and 'Tetramymena: Intorallelic repression. Timing of event. Series of genes for serotype: 14; only one active at a time; otiler Bluplic te" <;;enes turned off, Other genes: behave similarly. I'iming of events: controlled. Chees tAnrough environmental chan,es: the s?rotypes, 3. The position effect in Drosophila: Decker: white locus next to heterochromatin: timing and frequency of this. df:'ect of Y. Lioes not al-tt>r time of event; alters frequency among celis in which event occurs0 4. The pro&eins of the 7s comp.:nent of antibodies: amino acid differences in one segment of this compoilent. 5. Controlling elements: maize. Combonents that may be identified and chLiracterized; serve to mouifg gere action and at p.lrticulsr times and in particular manners0 6. The effects of hormones: act at gene level. Xffects only in certain cells. dombine with chrov!oso::ie - some evideiice of thi-, -2- III. The components of the chromoso ,es in nucleated organisms: 1. Enormous difference in associated components of LX&L betIdeen bacteria and organisms with chromosomes and nuclei. 2. This difference: undoubtedly associated with mechanisms of reproduction of the chromoso:i:e and with control of gene action diiring diffc?:entiation and in individual cells, ,alre;tdy difi'erentiat 3. Mrnber of L)X\jri molecules per chroliloso,se: few. a number oi them per chromosoi:le. Number of replicons: 4. The components of the chromos:;f:e: Histones: LycPne rich and arginine rich: Basidual protein: acidic the new R~LA species: with histones; with basic nrotein. Ihe phosphoproteins; the phospholipids. 5. The Histones: have been the candidate for repression of gene action: 4. nistones in :J.ctiVe and in ctive chromosor;ie parts: not didferent. dome turn-over of hi%ones without replic.zLtian of UiL~O b). ;"ih,jt his-tones might ,re doing: csescribe shortly; 6. The organization of parts of chromtisoi:es in the light microscope: (1). 'The nucleolus organizer: J'unction; cytoplasmic ribosomes, Slides 2, 2. Qtaiuq$ t)lfY&wdeh- 3 lLNL&J (2). I'he true heterochrc)mtitin in the chro~oso es: psotions. About centromeres in I)rosophila and other or;l;nisms. L-ippe3rance in the nucleus. .it ends of chro1:ios!.;mes ; iit speci:il regi: ns: knobs in m;lize, W. Yhe position of the diffe ent p,-lrts in the working nucleus: -3- (3). The extensi e studies of Calf thymus lymphocytes in Grsky laboratory, Rockefeller Institute, a>. The apnpe :rznce of the nuclei: Before and aftersqeiiing. Slide 8 ,' Glide go b). the position of the co:.densed pdrts with respect to the nucle:ir membrane, CL I'he tests of the active and the inactive chromatin: Xadioacti-;e uracil - positions of formation of Rl\i~ :iutoradiogra.ph: Slide 10. (pas lti.on of c Aen:ed ;d--I.rts j. d). 'ihe mode of connections 01 the parts in the co:;der,sed regi ,n: Gross linkages bet?: -en strdnds produced by 1,ycine rich his-tones: associ:ted with phosphoric acid group of DNa This histone associ<..ted with the repression process thrcugh maintaining the clumps. i-irginine rich his-tones: also associLl-Led with the phosphoric acid group of i>ih but combine along siae of i)i\i!hO (4). Each nucleus,has its own type of contraction of chromatin, involv&ng diffeA.ent parts of the chromosoiL;es-- diffe sent genes o &*Q+& h&.43 @44u+f, q(5). The major question: \?I' A,, A: how does the differential contro14"take place? rih:-it genetic compor,ents are involved in this? n (6). pqMt~~;;u.~!r A schedule'of differential se:iuen::es of condensations must be present from zygote stage on, if co~.densation is one of' the mechanisms of co;.trol of gene action, r&j) q' &. ,,,'rr*:rw j. '$I!. `: ',:'dtf with iz!; Are there special elements in the chro:,o:::res that are ssocited 3 iieturn to case of control of esterase alleles in maize: conLro1 "oi time of action is different for each allele. 'Lhe control of t?is for these alleles is associ3t,?d with some component at the locus of the gene itself, Controlling elements in maize: the::e a::e candidates for such elements that serve as co .trols of gene action: j)le/3lan$lru,~*~tiswf~~ "4P* (8). Do we h,ve other evidence of controlling elements or cris;rol regions within the chrimosol!:e'$%#&M W9.$? (a). the beh:lvior of the B-type chro.,iosome in maize at pollen division. Control of non-disjunction: 'ihe lot-tion of the signaler for this: at heterochr m tic end. The loc+tion of the s m the signals: near the centromere end of the chro 'osolleO Both must be present for non-disjunction to occur0 Q6.r >- .~i&@w~~:' tividence from rye chromosol:,e: B-tye here: similar to maize Two element sys-i;em of co:.trol of non-disjunctiJi.s, at specific stsge in development, (b) ihe relationshi elements. !! of stage to the effectiveness of controlling 'hi? condens;ltion. brings us to the Class II type of -4- CL&% II CONDENSATIONS: Condensation of consequtive regions: Yarts of a chro;:losome; whole chroT:oso81e; whole set of chrso Known to be inactive for genes. Ii~ipOjj~j:~4 T 1. Example: X chrot',osoiiie in mammals: non-contrActed XY, xx, xxx, xxxx, xxxx: Selection of X to be nr~ti=nrk&, if more than one present, ~tla,~& W&@&& kosition of X in nucleus: Zlide 11 :lypotheslEs: All 8 chromosumes donaitioned for contraction; one only is rescued from th$s, 2. The control region for the contraction: the responder to some signal, In one region of X chr.3:`.osome O The tests in mause: X xutosome transloc~~tions. Xesults: contraction to both sides of the control regionL+~~~~~*r:' males 3. Condensation of whole set of chroriiosomes: me:ily bugs:,the set bd+?m& p4 received from the father: Slide l& 2osition in nqrcieus, This set represents the set previously received from the grand- mother. titline: Females: P'emale producing; dame female, eggs l-Lid later: "'ale producing, ;,(*.~l& Set from father in females: euchrom::tic 4&$ Set from father in males: heterochromatic, `4 IIW'O.~1,i!.1;V T : Set from father: condktioned to condense; rescue from this occurs in eggs destined to produce females; no rescue from this if it is to produce a male. SAlpjll$j;iT y'H()&$s: Setting occurs in the germline of the father in the chromosome set it had received from its mother, Heterochromatic set it received from its f;tther is disc;$rded; does not get into sperm. 4. The relation between setting for heterochromatization and for elimination: uses the sizme mechanism: &I~ llap.u& some mealy bugs: Set from father is elimi q-%3 ted in early cleavage of cells destined to become soma cells, This important for my tnesis. V. The control of the elimination process: reaction of chro11:os~~es destined for elimination during cleavage to cytoplasmic substiince which rescues this: Cecidomyid. 1. 'l'he Egg: Pole plasm. The reticulate substance in pole plasm. dhe pole plasm and the germ line, a>. Normal beh;-jvior: b,>. To show tile rescue from elimination related to reticulate substance (but not the pole plasm@. Example of one type of test: Ligature: 4. Centri@.&ation studies: Any nucleus that comes adjacent to reticulate substance will have its E chroirlosomes rescued from elimination process0 VI. The tests of the res,;onder and the signaler for the elimination process in Sciara and the rescue mechanism, Its relation to control of ~&tixwacf;i~ condensation of only one X in mammals: the rescue from condensC-ttion of only one A, 1, Sciara germ line of male: later stages; l*leiosis; constitution of sperm, 2. The Females: Two types: X' x: -- kroduces only females, normally 2 &: Produces males only, ".&I To show that the signaler is'yn the X'and not the Xr and that the re&?-'?$'r ig in the-heterochom chronoso:ze one location in the X,; This component was set in male germ line: to bfgict elimination of both ii chromosomes during 7th or 8th division. Y-: $ The elir@nations : <' Eggs of x of X in gzsrxurkk=x soma of eggs produced by X females: only one of two sister X chrL)l:losomes from father -2:. II II X X Females: B&JC both E chr~;~3so;~les from f,ther eli$in.,ted. -6- 3. Relation of control of elimination in the soma cells to product of the X' chromosome: Females: X1 X== normal1 female produces: one A from male elimin*;ited, Non-disjunctions:at meiosis: No X Chru:!iOSOiLie in e#@ nucleus Zygote: 1A +2X1A doma elimination: One X only eliminated, -Female: XX= normal zygote 1X + lx Soma: 2X + 18, elimination of both i from father. 1iondisjunctions: 2 X chromosomes in egg from mother. zygote: 2iS+lA/2X+lA. "0th X chromosomes from male elimin ted. Hypothesis: 80th X chromosoxieg from father destined for eliminatitin in the soma. X chromosome produces some "particle" that is able to rescue only one X chromoso;,le . ilust be in cytoplasm as in non-disjunction case, no X from female in the zygotes. Setting: Occurs to element 1oc:Lted in het,,rochro:,l-itin of Xi: xxxx * v X- Autosome trxnsloc Ations: Any part of chromoso18,e com;?lement th:t carries this element will follow elimination path, non-disjunction path at meiosis of male. Best trxnsloc tion: X . . . . . " setting occurs during germ line of male to this elel,lent. Rescue occurs in soma of females to one eleinent No settings occur to this element, of female. leading to elimination in germ line Unce rescued, rr-maines rescudd until it a:.!in passes through germ line of male. 4. The rel;;tiun of the dciara c:~:;e to X chroi.losomes in mammals: lhe setting rcgion:for corLdensxtion, Only one X rescued. signaler not in ii chrorosone probL2bly. ahouid be in o:!e of the autosoxes. VII. The setting of the controlling elements in maize and the resetting process, -iVill discuss later,