MATERNAL-ZYGOTIC LETHAL INTERACTIONS IN DROSOPHILA MELANOGASTER: THE EFFECTS OF DEFICIENCIES IN THE

ZESTE-WHITE REGION OF THE X CHROMOSOME1

LEONARD G. ROBBINS

Genetics Program and Department of Zoology, Colleges of Natural ScLnces and Osteopathic Medicine, Michigan State University, East Lansing, Michigan 48824

Manuscript received April 12, 1980 Revised copy received June 16, 1980

ABSTRACT

The possibility that essential loci in the zeste-white region of the Dro- sophila mlanogaster X chromosome are expressed both maternally and zy- gotically has been tested. Maternal gene activity was varied by altering gene dose, and zygotic gene activity was manipulated by use of position-effect variegation of a duplication. Viability is affected when both maternal and zygotic gene activity are reduced, but not when either maternal or zygotic gene activity is normal. Tests of a set of overlapping deficiencies demonstrate that at least three sections of the zeste-white region yield maternal zygotic lethal interactions. Single-cistron mutations at two loci in one of these seg- ments have been tested, and maternal heterozygosity for mutations at both loci give lethal responses of mutant-duplication zygotes. Thus, at least four of the 13 essential functions coded in the zeste-white region are active both ma- ternally and zygotically, suggesting that a substantial fraction of the genome may function at both stages. The normal survival of zygotes when either maternal gene expression or zygotic gene expression is normal, and their in- viability when both are depressed, suggests that a developmental stage exists when maternally determined functions and zygotically coded functions are both in use.

HERE is substantial genetic and molecular evidence for the developmental importance of maternal gene action (for reviews see DAVIDSON 1976; KING

and MOHLER 1975). Are maternally active genes a restricted subset of genes? Do genes that determine maternal functions also determine zygotic functions? What fraction of the genome determines functions required both maternally and zygotically? There is some evidence that genes with both maternal and zygotic function are not uncommon. GALAU et al. (1976) have presented evidence that, in the sea urchin, the subsets of polysomal RNA information found at later embryonic stages are all included within the oocyte RNA sequences. RIPOLL (1977) and RIPOLL and GARCIA-BELLIDO (1979) have shown that a majority of Drosophila lethal mutants that are both cell-autonomous and cell-lethal never- theless survive till hatching of first-instar larvae. Their interpretation is that these embryos survive because maternally specified equivalent functions are

1 Research supported by National Science Foundation Grant PCM 79-01824.

Genetics 96: 187-200 September, 1980

188 L. G . ROBBINS

active during embryogenesis. In addition, GARCIA-BELLIDO and Moscoso DEL PRADO ( 1979) have suggested that maternal-zygotic functions are widespread since offspring of deficiency-heterozygous females often have reduced viability, while genetically identical embryos fathered by deficiency-heterozygous males do not. Whether the defect that kills off spring of deficiency-heterozygous mothers relatively early in embryogenesis is the same as that which kills deficiency- homozygous embryos at a later stage was not demonstrated, nor was it demon- strated that reduced maternal gene dose, rather than some property of the pater- nally transmitted normal homolog, causes the nonreciprocal lethality. That pa- ternally and maternally transmitted chromosomes differ has been demonstrated by the recovery of mutants that differentially affect the behavior of paternally and maternally derived chromosomes (BAKER 1975; HALL, GELBART and KAN- KEL 1976) and has been raised in various reports of parental effects on variega- tion ( SPOFFORD 1976).

A number of Drosophila genes are known that have both maternal and zy- gotic functions. In most instances these mutants were detected as the result of chance observations, and the conditions that expose the maternal-zygotic inter- actions are not of general utility for detecting maternal-zygotic interactions at other loci. Lethals, which are common and easily detected, would be the mu- tants of choice for asking about genes that act both maternally and zygotically but for the fact that an unconditional zygotic lethal cannot be tested for maternal effects, at least not as a homozygote. The recovery of temperature-sensitive ma- ternal and zygotic lethal mutations ( SHEARN, HERSPERGER and HERSPERGER 1978) offers one approach to uncovering these functions. Another approach, and one that is well suited to asking the general questions posed above, is to select a wel!-defined region of the genome in which all essential loci are known, where a large collection of lethal mutants already exists, and to devise a set of condi- tions that allows testing for maternal effects.

This approach is similar to that used by GARCIA-BELLIDO and Moscoso DEL PRADO (1979) in that maternal effects of deficiency-heterozygous females were initially examined. However, use of duplications corresponding to the deficien- cies, and modulation of gene activity in those duplications by position-eff ect variegation, allows demonstration that the maternal-effect lethality is, in fact, a consequence of reduced gene dose and is restricted to embryos that do not have fully active complements for the region. Demonstration of similar maternal- zygotic interactions for several single-cistron mutants confirms that individual gene functions are active during both oogenesis and zygotic development.

CROSSES A N D RESULTS

The deficiencies, duplications and zeste-white regioE mutations used in these experiments were supplied by B. JUDD. Descriptions may be found in LINDSLEY and GRELL (1968), JUDD, SHEN and KAUFMAN (1972), KAUFMAN et al. (1975) or SHANNON et al. (1972). Except as noted, descriptions of other markers and

MATERNAL-ZYGOTIC LETHAL INTERACTIONS 189

chromosomes used may be found in LINDSLEY and GRELL (1968). All crosses were done on cornmeal, molasses, brewers' yeast medium at 25", except as noted.

Exposing maternal-zygotic gene interactions: Homozygous females cannot be used to test lethal mutants for maternal effects. Use of heterozygous females re- quires exposing effects of reduced, rather than absent, maternal gene activity. For a gene that acts both maternally and zygotically, reduced maternal gene activity may be more apparent if the same gene function is depressed in the zygote as well. Thus, in order to determine whether the zeste-white interval contains essential functions that are expressed either maternally or zygotically, or both maternally and zygotically, the interaction between reduced levels of maternal and zygotic gene activity was examined. The results of these experi- ments are shown in Table 1.

Df(1) K95 and Df (1) ~ ~ ~ ~ - 4 ~ are two small, nonoverlapping deficiencies that between them delete most of the zeste-white interval. The level of maternal gene product was reduced by using females heterozygous for either of these deficien- cies. To reduce the level of zygotic gene activity, the crosses were done so as to generate a class of male zygotes that carried either one of the deficiencies and a small duplication. That duplication, Dp(j ;4)mg, is a segment of the X chro- mosome that includes the entire zeste-white region translocated to the fourth chromosome (ROBBINS 1977). Thus, Df/O ; Dp(1;4)mg/+ males are euploid but, as is demonstrated below, position-eff ect variegation of the duplication re- sults in less than normal gene activity. In the crosses shown in Table 1, defi- ciency males are generated in two ways: by crossing deficiency-heterozygous

TABLE 1

Maternal-zygotic lethal interactions of two zeste-white region deficiencies ~ ~~ ~

Maternal deficiency - Daughters Sons 1.0s~ of Recovery of

Cross Non-Df Df,Dp Non-D\ Df,Dp Y"X.YL Df,Dp sons

Father: X Y / O ; Dp/4 1. Mother: y p n 973 - 1329 - 0.27 - 2. Mother: K95/y 484 448 626 19' 0.26 0.06 3. Mother: w258/y 482 477 620 216 0.23 0.70

Paternal deficiency Mother: C ( l ) R M / O ; Dp/4

- - 4. Father: y p n 155 - 185 - 5. Father: K95/w+Y 483 - - 196 - 0.81 6. Father: w258/w+Y 309 - - 1 43 - 0.93

* May include some nondeficienry crossovers between y and the deficiency. == YSX.YL, y B. Dp = Dp(l;4)mg. 4,= spapol. K95 = Df( l )K95, yL. wz58 = Df(l)w258-45, y z w-. C(1)RM = C(I)RM,yS S U ( W ~ ) w5.

The deficiency males produced in these crosses are Df/O; Dp(l;#)mg/+. In the maternal transmission crosses, the frequency of females is depressed because of loss of the compound-XY in the male parent. That loss is calculated as 1 - (daughters/sons) for the control and as 1 - (daughters/2~non-Df sons) for the two deficiency crosses. Recovery of Df;Dp sons is cal- culated as ZXDf;Dp/non-Df sons to account for lethality of Df;non-Dp sons. In the paternal transmission crosses, recovery is similarly calculated as 2 x Df; Dp sons/daughters.

190 L. G. ROBBINS

females to attached-XY (= YsX.YL) males and by crossing deficiency-bearing males to attached-X [= C(Z)RM] females. Attention is first drawn to the Df (1 j K95 results and to the following observations: (1 ) Df;Dp males survive despite variegation of the duplication if their mothers are euploid (cross 5 ; re- covery = 0.81). Thus, there is only a slight effect of this level of reduced zy- gotic gene activity 011 survival; (2) the recovery of deficiency females, which also carry a normal X chromosome, and in half the zygotes the duplication as well, is unaffected by maternal deficiency heterozygosity. They are recovered as often as nondeficiency females (cross 2; 448 us. 484); and ( 3 ) there is a striking absence of Df(Z)K95 ; Dp(Z;4)mg males when the female parent is heterozygous for the deficiency (cross 2; recovery = 0.06). Thus, there appears to be a synergistic interaction between reduced maternal and reduced zygotic gene activity.

The D f ( 1 j ~ 9 ~ ~ - ~ ~ crosses also show a maternal effect that is exposed in Df;Dp sons, but not in deficiency-heterozygous daughters. Though less extreme, this effect is statistically significant (the difference from a 2: 1 expectation of normal : deficiency males yields x2 = 21.6, with 1 d.f.) and is replicable.

An objection might be raised to the notion that maternal-zygotic interaction lethality is more extreme than expected from separate maternal and zygotic ef- fects. Were it the case that maternal deficiency heterozygosity caused substan- tial inviability of all off spring classes. a weak zygotic effect coupled with a strong maternal effect could yield the apparent interaction. In the foregoing experi- ments, a maternal lethal effect on all offspring genotypes would not be detected except as a reduction in fecundity.

This matter has been addressed in the following way. Df(l)K95, y z / y females were again crossed to YsX.YL, y B/O ; Dp(l;4)mg/spaPoz males. Eggs were col- lected over a 24 hr period on yeasted, grape-juice-darkened medium. After count- ing, the eggs were collected and transferred to normal food and allowed to develop. Of the 2.094 eggs sampled, 804 survived as females and 541 as nonde- ficiency males. Recovery of 541 nondeficiency male zygotes implies that 541 deficiency male zygotes would have been expected. Thus, 1,886 out of 2,094, or 90%, of the eggs are accounted for as survivors, deficiency males, or as victims of the maternal-zygotic interaction. There is no room here for a maternal effect sufficient to account, in a multiplicative fashion, for the lethality of Df;Dp sons of deficiency-heterozygous mothers. I t may be noted that, though no attempt was made to define the particular stage at which the lethal zygotes succumb, very few survive to o r beyond the white-pupa stage.

These observations suggest that, for at least two segments of the zeste-white region: ( 1 ) a necessary oocyte function is impaired when gene dose is reduced in the female; (2) the zeste-white genes carried in Dp(l ;$ jmg have less than normal activity; and (3) combination of the maternal and zygotic defects, but neither defect alone, results in lethality.

Several experiments have provided substantial confirmation of the two major elements of this hypothesis: ( 1 ) the maternal effect is a consequence of reduced gene dose and not a consequence of a peculiar sex-specific response of the dupli-

MATERNAL-ZYGOTIC LETHAL INTERACTIONS 191

cation, and (2) the inability of Dp(1;4)mg to ensure survival of Df;Dp males is a consequence of reduced activity resulting from position-eff ect variegation of the duplication.

Gene dose and the maternal e&ct: Two experiments demonstrate that the maternal effect is a consequence of reduced gene dose. Since the effect of Df(l)K95 is more readily followed, that deficiency was used in these experi- ments. First, as shown in Table 2. paternal transmission of the duplication in the absence of a maternal deficiency does not result in reduced recovery of Df;Dp males. The recovery of Df;Dp males is not significantly different, when both the duplication and deGciency are paternally transmitted, from that ob- served in Table 1 when only the deficiency came from the father. The second cross of Table 2 provides an additional control where, as discussed in more de- tail later, variegation of Dp(l ;4)mg is suppressed. The difference in recovery is not statistically significant. Thus, the parental effect on survival is a maternal and not a paternal effect.

The second experiment tested whether the maternal effect was a consequence of reduced gene dose, and not a concequence of maternal transmission per se. To examine this, maternal gene dose was increased by including both the de- ficiency and duplication in the female parent’s gefiotype. Two such crosses were done: one where only the deficiency female parent carried the duplication and the other where both parents carried the duplication. For the latter cross, chro- mosome 4 markers allowed identification of progeny carrying a paternally de- rived duplication. Deficiency; nonduplication females were tested as a control. The results of these crosses are shown in Table 3.

It is apparent in these data that the presence of a duplication in the deficiency- heterozygous mothers substantially improves the viability of Df; Dp sons. As the second cross indicates, all of the expected Df;Dp males are recovered when their mothers carry a duplication as well as a deficiency; whereas, genotypically iden- tical sons of females lacking the duplication are recovered only 7% of the time. Furthermore, as seen in the third cross, rescue afforded by a maternal duplica- tion occurs even when off spring do not receive the maternal copy of the duplica- tion-47% of the Df;Dp males survive if they receive the duplication from their fathers. Even though recovery of sons bearing a paternal duplication is lower than that of sons bearing a maternal duplication, the calculated values cannot be taken too literally. The paternal-duplication-bearing males are ciD in pheno-

TABLE 2

Effect of paternal fransmissicn of Dp (1;4) m g on recovery of Df ;Dp males

C(1)RM daughters Dj;Dp sons Recovery __- Cross Df/Y; Dp/+ x C(I)RM/O; spapo~/spapo~ 724 254 0.7 Df /Y; D p / f x C ( l ) R M / Y ; spapo1/spapo1 200 93 0.9

Df = Df( l )K95, ys. Dp = Dp(l ;4)mg. Recoveries are calculated as 2 x Df;Dp males/females.

1 92 >. G . ROBBINS

MATERNAL-ZYGOTIC LETHAL INTERACTIONS 193

type and their viability is depressed for that reason alone. It may be noted that an equivalent disparity in the recovery of ciD and 4- is seen for both the deficiency and nondeficiency females, though ciD and 4- recovery is closer to equality among the nondeficiency males. In addition, though Dp(l;#)mg is homozygous lethal in otherwise normal flies, Df ( I ) K 9 5 / 0 ; Dp(l ;4)mg/Dp( l ;4)mg males have been found (in other crosses) occasionally to survive. Thus, the number of males apparently carrying only maternally transmitted duplications is inflated by survival of an unknown number of males that carry both maternally and pa- ternally derived duplications, while the number of males carrying a paternally derived duplication is deflated by the effect of ciD on viability. Even if some of the apparent numerical discrepancy reflects an effect of parental origin on variegation (SPOFFORD 1976), it is neverthless clear that normal gene dose in the female, though probably still providing less than normal gene activity, sub- stantially restores the viability of Df;Dp sons. Together, these data establish that the maternal effect observed for deficiency heterozygotes is an effect of reduced gene dose and not some peculiarity of maternal transmission itself.

Variegation and reduced zygotic g e m activity: The second major element of the hypothesis is that Dp(l;#)mg is less than fully active. Position-effect variega- tion (reviewed by SPOFFORD 1976) is a common consequence of chromosome rearrangements and was tested for by inquiring whether the viability of Df;Dp males responds to agents known to affect variegation. Two responses were tested: relaxation of variegation by addition of a Y chromosome and enhancement by reduced temperature. The effect of a Y chromosome on survival of Df(l)K95 and D f ( l ) ~ d ~ ~ - ~ ~ deficiency; duplication sons of deficiency-heterozygous females was tested at 25", survival of Df ( l ) K 9 5 / 0 ; Dp(1;4)mg/spdo2 males was tested at 19", 25" and 29", and both the maternal effect and Y chromosome effect were checked for at 19". In addition, the behavior of a different dupli- cation, visibly variegating for an included white locus, was examined. The ex- pected enhancement of viability by addition of a Y chromosome is clearly shown in Table 4.

The effect of temperature on the survival of Df( l )K95 /0 j Dp(l;4)mg/sp.aPo2 males is shown in Table 5. The most common response of variegating systems to

TABLE 4

Suppression of lethality of D f ;Dp sons b y a Y chromosome

Non-Df D f Non-Df Dp;Df sons Recovery Mother Father daughters daughters sons

- XY/O; Dp/4 1551 1523 1927 62 0.12 - of (lY{959/Y X Y / Y ; Dp/4 805 763 822 225 0.55

Df(i)w"8-45/y XY/O; Dp/4 2262 2245 281 1 1068 0.76 - - X Y / Y ; Dp/4 548 542 515 234 0.19

xy=YSX.YL, y B, Dp=Dp(Z;4)mg. Df(l)K95=Df(l)K95, y2. Df(l)w258-45=Df(l)wzS8-4s, yg w-, Recovery of Df;Dp sons is calculated as 2 ~ D f ; D p sons/non-Df sons.

1941 L. G . ROBBINS

TABLE 5

The effect of temperature on gene expression in Dp (1 ;4)mg

Kon-DI; of; Non-Df; Non-Dp Non-Df, Dp Non-Dp DJ;Dp Non-Dp Kon-Df,Dp DI,DP

sons sons -

lemperatuie daughters daughters daughtas daughters sons

29" 385 261 (0.68) 411 437 510 297 (0.58) 19 (0.04) 2 5 O 959 656 (0.68) 833 808 1287 958 (0.74) 43 (0.03) 19" 1079 934 (0.87) 912 958 1650 1343 (0.91) 7 (0.004)

Homogeneity xz, 2 d.f. 38.7 17.0 14.8

Df(l)Ii95,. y2/y; spap*l/spal.lol females were crossed to Y S X YL, y B/O; Dp(l ;4)mg/~pa~ '~~

Non-Df; Dp daughters: non-Df; Dp daughters/non-Df; non-Dp daughters. Non-Df;Dp sons: non-Df;Dp sons/non-Df; non-Dp sons. Df;Dp sons: Df;Dp sons/non-Df; non-Dp sons. Each group of females tested was also subdivided by whether they were themselves reared at

29", 25" or 19". However, there was no grandparental temperature effect and those homogeneous results have been combined.

males at the indicated temperatures. Recoveries (in parentheses) were calculated as follows:

temperature shifts is reduced gene expression at lower temperatures. Survival of Df;Dp males is therefore expected to decrease, and does decrease, as temperature is reduced. In addition, hyperploidy for Dp(I;4)mg also causes some loss of viability. One therefore expects, and finds, increased survival of hyperploids at lower temperatures. These observations are all in accord with reduced activity of Dp(I;4)mg at reduced temperature and are therefore in accord with the idea that Dp(I;4)mg variegates for the functions it encodes.

Df ( I )wP58-45 yields only a weak maternal-zygotic interaction at 25". If its effect is analogous to the more striking effect of D f ( l ) K 9 5 , it should be possible to enhance it by increasing variegatioE of the duplication. Reduced temperature accomplishes this, as indicated by the data in Table 6. While survival of Df ( I ) ; Dp(I;4)mg/spaPo1 sons of deficiency-heterozygous females is about 70% at 25" (e . g., Table 7, cross 3 ) , it is reduced to only 10% at 19" (Table 7, cross 2). As at 25", survival is markedly improved by addition of a Y chromosome (Table 7, cross 2 us. Table 7, cross 4).

TABLE 6

Suruiual of Df (1) wz58--45; Dp(l;4)mg males and effects of a Y chromosome at 29" ~

Daughters Sons Recovery of Cross Father Mother Non-Df Df Non-Df D f Df;Dp sans

1. Df/wfY C(I )RM/O; Dp/4 1228 - - 6 74 1.10 2. m/O; Dp/4 Df/Y 1192 1172 1768 85 0.10 3. Df/w+Y C(I)DX/Y; Dp/4 1588 - - 948 1.19 4. B / Y ; Dp/4 Df/Y 559 635 710 203 0.57

Df =Df(2)~"8-4~, y2 W-. X Y =YsX.YL, y B . C(I )RM=C( l )RM, y2 S U ( W ~ ) @. C(Z)DX=

Calculation of recovery of Df sons is as indicated in Table 1. C(I)DX, y f . 4 ~ = s p a p o ~ .

MATERNAL-ZYGOTIC L E T H A L I N T E R A C T I O N S 195

While the foregoing experiments demonstrate that the inability of Dp(1;4) mg to rescue deficiency zygotes is a consequence of variegation, it remains possible that variegation is a necessary, but not sufficient, explanation for the behavior of Dp(1;4)mg. The entire phenomenon could be peculiar to this particular dupli- cation. Whether other variegating duplications could also expose maternal- zygotic lethal interactions has been examined by using D p ( i ' ; 4 ) ~ " ~ ~ g in lieu of Dp(l;4)mg. Dp(1;4) wms5g includes only part of the zeste-white region, but does extend beyond the segment missing in D f ( l ) ~ ~ ~ * - ~ ~ . Unlike the situation with Dp(l;4)mg, variegation of D ~ ( 1 ; 4 ) w " ~ ~ g is quite apparent; w/O ; Dp- (l;4)wnTfi5g/+ males have eyes that are almost entirely white. Addition of a Y chromosome produces an obvious increase in the amount of red tissue, but still leaves most of the eye white. Since Z I ~ ( I ; ~ ) W ~ ~ ~ ~ ~ ~ shows little gene activity in normal circumstances, tests of its ability to rescue Df ( I ) w " * - ~ ~ zygotes were done at 29" to reduce variegation. Survival was examined both in the presence and absence of a Y chromosome. The results of these crosses are shown in Table 7.

In the absence of a Y chromosome, Df ( I ) U ? J * - ~ ~ ; Dp(2;4) wmfi5g zygotes do not survive whether the deficiency is maternally or paternally transmitted (Table 8, crosses 1 and 2). However, when Df/Y ; Dp/+ males are generated, a maternal effect on survival is evident. Only 20% of Df/Y;Dp sons of deficiency females are recovered (Table 8, cross 4), although recovery of identical males from normal mothers is near normal (Table 8, cross 3) . Furthermore, survival is much reduced here as compared with the approximately 70% survival observed when Dp(1;4)mg is used. Thus, a weak maternal-zygotic lethal interaction is even more clearly exposed by Dp(l;)wnTfi5g than by Dp(1;4)mg.

The generality of maternul-zygotic lethal interactions: The preceding results indicate that genes that function both maternally and zygotically can be de- tected by combination of reduced maternal and reduced zygotic gene activities. How common are genes that function at both times? As a first step in examining this, a set of overlapping deficiencies that dissect the zeste-white region have been tested for maternal effects on survival of Df;Dp zygotes. In each case, Df;Dp zygotes were generated by transmission of the deficiency from the mother in one cross and from the father in a second cross. As a consequence of previous

TABLE 7

Maternal-zygotic lethaal interactions of Df (1) w258-45 and Dp (1 ;4) wm65g

Daughters Sons Recovery of Cross Mother Father Non-Df D f Non-Dj Df;Dp Df ;Dp sons

1. C(I )RM/O; Dp/4 Df/w+Y 2776 - - 0 0.0 2. Df/Y X Y/O; Dp/4 1514 1528 2083 0 0.0

4. Df/Y X Y / Y ; Dp/4 1212 1214 1429 141 0.20

__

3. C(Z)DX/Y; Dp/4 Df/w+Y 1824 - - 883 0.97 __

Df=Df(i)wZ-45, y2 W-. Dp=Dp(l;4)wm659. C(I )RM=C(I )RM, yz su(wa) @. C ( I ) D X =

Crosses were done at 29". Calculation of recofvery of deficiency males is as indicated in Table 1. C ( I ) D X , y f . X Y YSX.YL, y B.

196 L. G . ROBBINS

TABLE 8

Maternal-zygotic lethal interactions of zeste-white region deficiencies

Female transmission- Non-Df D f Noil-Df Df:Dp

Deficiency tested daughters daughters sons sons Recovery

Male transmission _ - _ _ Non-Df DkDu

daughters ions Recovery

Df (1) wrJ1, y2 Df(1)64c4; y+ DI ( 1 ) w258-11 ,Y Df (1)64j4, y Df(l)wX12, y Df (1)65j26, y + Df (1 ) 62gl8, yf

Df(1)64fl, yf Df (1)K95, y2

Df ( 1 ) W'JZ, y+

Df (1) ~ 2 5 8 - 4 5 , YZ

209 1116 75 1 92 1 1141 1260 1186 1111 1171 1262 968

219 1102 765 947 1137 1427 1228 1144 1148 1216 973

347 1399 1024 1102 1579 1671 1470 1419 1388 1466 1273

1 2 2 11 3

813 696 7

384 25 467

0.006 0.003 0.004 0.019 0.004 0.973 0.947 0.010 0.553 0.034 0.734

302 629 466 278 1431 820 512 740 469 214 158

146 0.97 370 1.18 55 0.24 211 1.52 388 0.54

315 1.23 343 0.93 306 1.30 126 1.18 85 1.08

4344 1.06

Ratio __ 0.01 0.00 0.02 0.01 0.01 0.92 0.77 0.01 0.42 0.03 0.68

Recovery of Df/Dp sons following maternal transmission of the deficiency was measured in crosses of either: (1) Df/y females, where the deficiency chromosome was marked by yf or y2; or (2) Df/y+ females, where the deficiency chromosome was marked by y . In each case, the females were crossed to YsX.YL, y B/O; Dp(i;4)mg/spaPoz males and recovery is calculated as 2 x Df;Dp sons/non-Df sons. Male transmission crosses were Df/w+Y males X C(I )RM, y2 su(wa) &/O; Dp(l;4)mg/spapol females and recovery is calculated at 2 x Df sons/non-Df daughters. The last column gives the ratio of recovery in the female transmission cross to that in the male transmission cross.

work (ROBBINS 1980), most of the X chromosome proximal to Df(l)K95 and and most autosomal material had been replaced with a common

wild-type background. The other deficiencies do not share a common background and some extraneous viability differences might be expected. Nevertheless, any nonmaternally determined viability differences would be exposed in both re-

gt tko z z w l zw8 zw4 zwl0 zw13 zw2 ZW3 zw6 zw12 zw7 zw5 zwll zw9 w

b D f ( I ) 6 2 g l 8 3 &f(1)64j4-+ L D f ( l ) w 2 5 8 - 4 5 - 68

- ---------------- '.42Df W64f I+

D f ( l )wrJ2 +Of (Il65j26- , .01 I I Df(l)wrJ1 .01

k Df(1)64c4 .oo

.02

I

I

4 I Df ( l)w258-1 I

Df ( l )wXl2 I

.03 - Df (I) K95- FIGURE 1 .-Deficiency map of maternal-zygotic lethal interactions. The relative recoveries

of Df; Dp males following maternal .versus paternal transmission of the deficiency are indicated, together with the arrangement of the deficiencies with respect to the lethal and visible loci in the zeste-white region. This figure is based on the data in Table 8.

MATERNAL-ZYGOTIC LETHAL INTERACTIONS 197

ciprocal crosses. Thus, attention may be focused on maternal effects by exami- nation of the ratio of recovery in the two crosses. All of these crosses were done at 25". The results are shown in Table 8 and are summarized diagrammatically in Figure 1.

There is no single segment of the zeste-white region that entirely accounts for the maternal effect, though there are substantial regional differences. The gt to z w l segment, defined by Df(l)62g18 and Df(l)65j26, evinces very little, and possibly no, maternal-effect lethality. On the other hand, the zwl3-w seg- ment, defined by Df(l)64j4, Df ( l )64f l and Df ( 1 ) ~ ~ ~ ~ - ~ ~ must contain at least three dosage-sensitive, maternally and zygotically acting sites. Heterozygosity for D f ( l ) ~ $ ~ ~ + ~ produces a relative recovery of 0.68; deficiency for the zw3- zw6 segment, included in the segment of Df( l )64f l that is not overlapped by Df further reduces recovery to 0.42; and maternal deficiency for the zw13-zw2 segment nearly eliminates survival of Df;Dp sons. Whether any sites within the zw8-zwlO segment have separate maternal effects cannot be deter- mined, since the effect of zw13-zw2 deficiency heterozygosity is so severe that any additional effects of the longer deficiencies would not be resolvable.

While exhaustive testing of zeste-white region single-gene mutations for maternal-zygotic lethal interactions is presently underway, initial observations indicate that the maternal effect described in this report is not restricted to de- letions, but can be ascribed to individual gene functions. Futhermore, these tests indicate that the deficiency map underestimates the number of maternally and zygotically active functions. Data for two zw2 mutations and three zw13 muta- tions are shown in Table 9. The maternal effect is quite as striking here as it is for deficiencies. The zeste-white region, therefore, encodes at least four, and conceivably more, functions that are both maternally and zygotically active.

TABLE 9

Maternal-zygotic interactions of zw2 and zw13 mutmts ~~ ~~ ~ _ _ _ _ ~ ~ ~ ~ ~ ~

Recovery of Mutant Daughters sons mutant; D p

Locus Allele parent Non-mutant Mutant Nor-mutant Mutant sons Ratio

6 2 ~ 2 1 Female 1300 1310 1798 71 0.08 0.08

65123 Female 948 1031 1462 57 0.08 0.07 zwz Male 1296 - - 6 73 I .04

Male 705 - - 403 1.14

64bl l Female 1814 1667 21 79 119 0.11 0.11

zw13 e50 Female 464 487 756 187 0.49 0.44

13 Female 631 585 838 42 0.10 0.10

Male 95 0 - - 472 0.99

Male 760 - - 420 1.11 Male 492 - - 248 1.01

Lethal/y females were crossed to YSX.YL, y B/O; Dp(l;4)mg/spaPOz males and lethal/w+Y males were crossed to C ( I ) R M , ye su(wU) Wa/O; Dp(l;4)mg/spaPo1 females. Calculations were done as indicated in Table 8.

198 L. G . ROBBINS

SHANNON et al. (1972) have reported on the lethality patterns of I2 of the 13 known essential zeste-white loci, including that of zw2. Flies hemizygous for zw2 alleles do not die before the embryonic-larval transition. Thus, an individual gene function that, when mutant results in lethality at the end of embryogenesis, is nevertheless active during oogenesis.

DISCUSSION

The combination of matercal mutant heterozygosity with zygotic gene varie- gation provides a system for the detection and analysis of genes that act both maternally and zygotically. It has the advantage, over de nouo searches for con- ditional mutations, of allowing use of mutations in segments of the genome that have already been well characterized. It has the disadvantage of being restricted to segments for which appropriate duplications are available. Nevertheless, that this circumstance provides conditional expression of a lethal phenotype suggests that the effects of maternal and zygotic gene expression must temporally over- lap. If an early nonlethal effect of maternal insufficiency were combined with a later nonlethal effect of zygotic insufficiency, survival would be expected. How- ever, the combination of maternal and zygotic defects can be lethal. While it is conceivable that this temporal overlap reflects the existence of relatively long- lived maternal messages (GERASIMOVA and SMIRNOVA 1979), much less direct functional equivalence is also possible. These results imply only that the ulti- mate effects of maternal and zygotic gene activity can at some point substitute for one another. The Df;Dp or mutant;Dp condition should be amenable to clonal analysis to define the timing of this interaction. For example, for a cell- autollomous deficiency, Df ;Dp clones in off spring of deficiency heterozygous fe- males should not survive during those stages where maternally determined func- tion is still needed, but would survive after that time.

There are not many essential genes in the Drosophila genome that act only maternally (ZALOKAR, AUDIT and ERK 1975). There may, however, be quite a few that act both maternally and zygotically. The deficiencies used in the present analysis divide the zeste-white region into 10 segments, many just one chro- momere in length. Several divisions, however, are defined only by relatively long deficiencies that cause such severe maternal-zygotic lethality that distinc- tions among them are impossible. Nevertheless, three distinct sites have been identified by the deficiencies that contain functions that are active both ma- ternally and zygotically. In addition, tests of the first two single-gene functions examined have shown that both cause maternal-zygotic lethality. If the zeste- white region is not atypical, and there is no a priori reason to suppose that it is, a substantial fraction, and conceivably most, of the genes of Drosophila are ac- tive both during oogenesis and during zygotic development. It is therefore pos- sible that most maternally determined embryonic functions are not different from the ordinary housekeeping functions that are required in any cell. If there are maternal messages that are of special significance in early development, they may be but a small fraction of all maternally coded information. That the

MATERNAL-ZYGOTIC LETHAL INTERACTIONS 199

observation of a larval lethal effective phase cannot be taken to imply restriction of gene activity to that particular stage has already been noted by SHEARN, HER- SPERGER and HERSPERGER (1978) and by GARCIA-BELLIDO and Moscoso DEL PRADO (1979) ; it is explicitly demonstrated here for two single-gene functions. The relationship between lethal effective phase and time of gene expression need not be straightforward. The temptation to conclude that genes that act ma- ternally are of specific importance during embryogenesis must be avoided as well; zygotic effects may not be apparent unless the maternal genome is also defective. These results, along with those of GARCIA-BELLIDO and Moscoso DEL PRADO ( 1 979), RIPOLL (1977) and RIPOLL and GARCIA-BELLIDO (1 979), suggest that caution should be exercised in the interpretation of phenogenetic observa- tions, whether the interpretntion be developmental or in terms of gene organiza- tion. This note of caution may be of particular importance when duplications are used to define cytological location or time of gene expression. It is possible that, without reciprocal crosses and controls for variegation, the results may be misleading.

appreciated.

KHESIN, R. B., 1947

NANCY VEENSTRA assisted the author in doing these experiments and her help is gratefully

Dr. A. GARC~A-BELLIDO has recently brought the following references to my attention: Maternal effect in Drosophila melanogaster. Spreading of maternal effect.

Doklady Acad. Nauk SSSR 58: 667-671. -- 1948a Maternal effect in DrosophiZa melanogaster. Influence of maternal genotype upon the rate of progeny’s development. Doklady Acad. Nauk SSSR 59: 561-564.. - 1948b Duration of the influence of maternal genotype upon the development of Drosophila mehogaster progeny. Doklady Acad. Nauk SSSR 59: 751-754.

I t is with pleasure that I note this earlier work that in part, parallels my work, as well as that of the Madrid group.

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Corresponding editor: T. C. KAUFMAN

ZALOKAR, M., C. AUDIT and J. ERK, 1975