Donald Forsha Jones (1928)
POLLEN-TUBE FACTORS (p. 20-34)
In maize, two types of endosperm, sweet and waxy, differing from normal endosperm, both segregate in deficient numbers in some crosses. Both of these endosperm characters represent a somewhat abnormal or defective type of storage material, and such seeds germinate less vigorously and the seedlings make a somewhat smaller growth. Another endosperm character of a more decided abnormal nature, one of the numerous defective seeds, also manifests itself in deficient numbers as shown by Mangelsdorf and Jones (1926). In rice, Parnell (1921) describes an endosperm type, called glutinous, which gives less recessives than expected in the second generation from a cross with the normal type. Similarly in peas, Sirks (1923) finds that in certain crosses the wrinkled type, as in maize, gives the same result.
The fact that all of these somewhat abnormal endosperm types which, compared with the normal type, carry a lower carbohydrate reserve, give deficient ratios might indicate that the factors for the abnormal endosperm themselves are responsible for the reduced numbers in which the characters reappear after crossing with normal. Brink (1925) and Brink and Burnham (1927) have given evidence to show that maize plants segregating for waxy seeds which give fewer recessives than expected show a still greater deficiency when the plants are homozygous for the sugary factor. The presence of this factor in the homozygous condition in the sporophyte in some way handicaps the waxy-carrying pollen tubes. The proportion of waxy seeds in the starchy and sugary seeds produced by plants segregating for both factors is the same, so that there is no direct effect of the sugary gene in the gametophyte; but the gametes produced by a recessive sugary plant are affected. The cytoplasm laid down by the sporophyte regulates the behavior of the gametes produced by it.
Additional evidence shows that the difference in the pollen tubes carrying the factors for abnormal endosperm involves more than the nature of the storage material in the endosperm. In the first place it should be noted that only in crosses with one type of starchy maize has the sugary endosperm given a deficient segregation. All crosses with other types have regularly given normal segregations. Also, other abnormal endosperm characters which store much less material than waxy endosperm segregate normally. Hutchison (1921) describes a "shrunken" endosperm which gives 25 percent recessives when crossed with normal types. Mangelsdorf (1926) has arranged these endosperm characters on a scale of defectiveness by comparing their growth curves with normal seeds on the same ears as shown in Figure 5. Waxy endosperm shows the least reduction from normal development as measured by the comparative weight of dry matter. Shrunken and sugary are about 10 percent reduced from normal. Many lethal and semilethal endosperm factors described by Mangelsdorf bring about a very marked reduction in storage material and yet segregate normally. For example, de13 shows a development of only 5 percent of normal, yet counts of over 1,000 seeds have given 3:1 segregation that does not differ significantly from expectation.
Fig. 5.—The relative development of various endosperm characters represented by points on the growth curve of normal seeds of maize. (From P. C. Mangelsdorf in Connecticut Agricultural Experiment Station Bulletin No. 279.)
On the other hand, other characters which do not affect the storage material in the seed also segregate in markedly deficient numbers. Dwarf plants in barley, described by Hallqvist and noted in the previous chapter, furnish a good example. The deviating segregations in Oenothera, Melandrium, Datura, and Pisum may also be cited in this connection. Coulter (1925) has also reported deficient segregation in aleurone color factors in maize in material which does not involve differences in endosperm composition.
Sirks (1923), in commenting on the peculiar ratios obtained in the wrinkled- and smooth-seeded peas, assumes a factor which influences the growth of the pollen tube. In his case a gene which speeds up the growth is linked with a determiner for smooth seed, causing the gametes carrying this endosperm-determiner to be presented to the ovules in greater numbers. This endosperm character is also linked with a factor affecting the tendrils. The same pollen-tube factor should also modify the ratios of tendriled and acacia plants; and this it does in some cases, giving sometimes a significant excess of recessives, and at other times a result which is in close agreement with a 3:1 ratio. The data as summarized by Wellensiek (1925) are given in Table IV. In this case the pollen-tube factor, if such there is, must be linked with the recessive gene for acacia leaves, since these are in greater numbers than expected. Deviations of more than 3 times the probable error above and below 25 percent are expected by random assortment, but not so large as in these cases and not all in one direction.
SEGREGATION OF THE TENDRIL FACTOR IN PEAS
|AUTHOR||NUMBER OF PLANTS||DEVIATIONS
|Vilmorin and Bateson||322||127||+14.75||2.38|
Evidence for gametophyte factors which modify the growth-rate of pollen tubes is found in segregations which are in some cases above and in other cases below the expected percentage of recessives, as well as others that are normal in different progenies from the same stock. This also indicates that the abnormal endosperm types themselves cannot be responsible for the deviations, since in one case the gametes which carry their genes take part less numerously in fertilization and in other cases more numerously.
Emerson (1924) found such a situation in the starchy-sugary factors in maize. The cross of rice pop corn as a starchy type with sugary gives a low-sugary ratio in F2 with about 15 percent of recessives. The starchy seeds from such ears in the next generation give some segregating progenies that are again low in recessives, some normal (25 percent), and a few high-sugary (35 percent). When the extracted recessives from low and normal segregating ears are backcrossed with the F1, the combinations normal x F1, F1 x normal, and F1 x low gave approximately a 50-50 segregation; but low x F1 showed only about 30 percent of recessives. Low-sugar strains by rice pop corn gave 25 percent recessives in F2; while low-sugar x other starchy types gave an excess of recessives in F2. These results are interpreted as due to a factor from rice pop corn, linked with the starchy-sugary genes, which influences the rate of pollen-tube growth.
Pollen-tube factors have also been found to be linked with another endosperm factor, one of an extensive series of defective seeds studied by P. C. Mangelsdorf (1926). The details have been published by Mangelsdorf and Jones (1926). Combined with Emerson's findings, they give a convincing evidence of gametophyte factors influencing the rate of pollen-tube growth and in that way affecting recombination.
A strain of maize segregating for defective seeds of the type described as de1 was crossed with 5 different and unrelated stocks. Fourteen segregating progenies were obtained with a total of over 5,000 seeds which gave 27.7 percent recessives. The deviation from 25 percent is 6.8 times the probable error and would occur by chance alone only once in several hundred thousand trials. Seven of these 14 ears show no significant deviations, 1 shows a deficiency, and the other 6 an excess greater than 3 times the probable error. These high-segregating ears combined give 31.9 percent recessives, the normal 25.1 and the low 20.0. These high and low segregations are not chance deviations otherwise they would give normal segregations in F3.
From 2 ears giving 32.4 and 33.6 recessives in F2, 18 segregating F3 progenies were grown, of which 6 are high with 32 percent recessives, 4 low with 17 percent, and 8 normal with 25.2 percent. The average for all lots combined is 26.4 percent, which is close to 25 percent. But the deviations from normal ratios of 10 of the 18 lots cannot be due to chance. Faulty classification is ruled out since errors of this kind would be in one direction. Other defective seed types have shown no such wide fluctuation in the percentage of recessives.
Evidence as to possible differences in the rate of pollen-tube growth in the plants segregating for defectives was obtained by separating the ears into upper and lower halves. The faster-growing tubes would be expected to furnish the gametes for fertilization of a greater number of seeds in the lower than in the upper halves on the supposition that the farther they had to travel the greater certainty that the faster individuals would reach the goal first.
In the segregating progenies which showed an excess of recessives there were 30.6 percent in the upper halves and 33 in the lower halves of the same ears. The normally segregating ears from the same parentage showed no appreciable differences in the two parts.
The difference in rate of pollen-tube growth is not due to the defective seed factor itself, since the deviations do not occur regularly in one direction. In some cases there are marked plus deviations, in others minus, while the others segregate normally.
The only way in which these peculiar results can be understood is on the assumption of a transmissible factor, situated in the same chromosome, that influences the rate of pollen-tube growth. Such a hypothetical gene has its expression in the gametophyte generation and has been given the factor symbol Ga. It is assumed to speed up the rate of growth in the pollen tubes which carry it. In the original stock in which the peculiar ratios were obtained it was linked with de1. For this reason the pollen tubes carrying de1 grew faster and a large proportion of them took part in fertilization and gave an excess of recessives.
The detailed workings of this factor and the actual results obtained have been reported elsewhere (Mangelsdorf and Jones, 1926). It should be noted here that only those plants which are heterozygous for the pollen-tube factor as well as the defective seed factor will give distorted ratios, and the direction of the deviation will depend upon the way in which the factors are linked. A plus-segregating plant will give in the next generation plus, minus, and normal segregations; and the proportions of these will depend upon two variables: the amount of crossing over and the ratio in which the Ga and ga are presented at fertilization.
While the results indicate that the factor which causes a deviation from normal segregation influences the rate of pollen-tube growth, the same effect maybe brought about in other ways. One possibility is that the presence of this factor in the heterozygous condition may cause a difference in germination of the pollen grains. Brink and Burnham (1927) believe that the pollen tubes carrying the waxy gene grow at a slower rate until the energy stored in the pollen grain is used up. Afterward, when the pollen tubes feed upon the stylar tissue, they grow at the same rate.
The close similarity of these results from defective seeds with those from sugary endosperm previously described has heightened interest from the fact that these two factors are situated in the same chromosome. Apparently the distortion in the ratios when rice pop corn is crossed with sweet endosperm is due to the same gametophyte factor present in the starchy parent and linked with the dominant Su endosperm factor. Since distorted ratios were obtained only when the F1 plants were self-pollinated or pollinated on to the dominant parent, it is evident that the gametophyte factor operates only when the same factor is present in. the sporophytic tissue of the seed parent. It apparently functions as well in the heterozygous as in the homozygous condition, and for that reason is considered as a dominant factor, although, when operating in the haploid condition in the male gametophyte, dominance would have no significance.
The crossing over between this defective seed factor de1 and sweet, su, using material which was free from the gametophyte factor, has been tested and found to be approximately 39 percent.
Since these two factors are linked and both are affected in the same way by a gene located on the same chromosome, the proof is reasonably complete that it is the same factor that is operating in both cases. On this assumption it is possible to estimate approximately the position on the chromosome of the gametophyte factor in relation to su and de1and at the same time determine roughly the ratio of Ga and ga gametes which accomplish fertilization.
This gametophyte factor which influences the rate of pollen-tube growth has been placed 23.2 units from Su and 27.0 units from de1 giving 50.2 units, which is approximately the map distance between Su and de1. With this location, and assuming that the gametes carrying the Ga factor accomplish fertilization 4.1 times as often as those which do not have it, the deviation of 7.8 percent in the defective seed ratio and of 8.8 percent in the sugary ratios are both in agreement with expectation. The relative effectiveness of the gametophyte factor in speeding up pollen-tube growth probably varies according to the length of the styles, weather conditions, and perhaps other influences.
From these calculations it is possible to predict the deviations from normal segregation which will occur when other characters on the same chromosome are crossed with types with which this gametophyte factor is involved. Wentz (1925) describes a defective seed type which is linked with sugary endosperm with 3.2 percent of crossing over. The crossing over between this factor and the gametophyte factor should be about 23.8 or 18.6 percent, depending on which side of the sugary locus it is situated. A cross between rice pop corn, which carries the gametophyte factor, and this defective seed type should give either 17.1 or 15.6 percent of recessives. One such cross made by Wentz actually gave 19.8 percent instead of 25 percent, as was the case in all the other crosses with other starchy types; and, considering only those ears that gave a significant deficiency, the average was 17.3 percent.
A similar situation undoubtedly exists in the deficient segregation of waxy endosperm in maize. Collins and Kempton first reported such segregation in 1911. Kempton (I9I9) has made an extensive investigation of the segregation of this endosperm character. Kiesselbach and Petersen (1926) have obtained additional data and have summarized all of the available figures to show that there is a deficiency of 1.1 percent from the expected 25 percent when the heterozygote is selfed and 0.7 percent from the expected 50 percent when the heterozygote is backcrossed to the recessive. They conclude, however, that "the evidence at hand seems insufficient to definitely establish the causes or significance of the deviations," although the deviations in each case are many times more than their probable errors and are certainly significant.
The difficulty of understanding the waxy segregations has not been due to any dearth of data but to the fact that all of the results have been lumped together, obscuring the situation in which some of the segregations have been in excess and others in deficiency. This is shown clearly by an analysis of Kempton's data obtained from a number of F1 plants that were intercrossed and also self-pollinated. By dividing these plants arbitrarily, according to their segregation when self-pollinated, into normal and low, using a deviation of twice the probable error as the basis of separation, the intercrosses can be arranged into the following groups: low x low, normal x low, low x normal, and normal x normal. If the deficiencies of some of the self-pollinated plants are mere chance deviations, then the different intercrosses, based on a classification of the self-pollinated plants, would give the same results. But actually plants, giving low ratios when selfed, give the same ratios when crossed on other plants. Normal plants give normal ratios when intercrossed. The conclusion is that the random assortment of the male gametes at fertilization is disturbed in about half of the plants and not in the other half. As Wellensiek, writing in English, well says: "It appears that adding results of various experimentists, which don't allow severally of drawing unambiguous conclusions, may be very hazardous"!
Kempton's results can be understood on the assumption that a gametophyte factor is involved similar to that which disturbs the sugary and defective seed segregations in another chromosome and is linked with the factor for waxy endosperm. Since a deficiency is noted in almost every case that waxy is crossed with a large number of other stocks, this factor must be associated with the waxy gene and work in the opposite direction to the factor which causes an excess of normal seeds in the rice pop crosses. It also is independent of the genetic constitution of the sporophytic tissues in which the pollen tubes grow, since the F2 plants backcrossed to the recessive give less than the expected number of waxy seeds. It thus differs in two ways from the sugary segregation. This factor probably affects the rate of pollen-tube growth, as in the other case. Brink (1925) finds that fewer waxy seeds are produced on inflorescences with long styles compared to those in which the styles were cut short.
Brink has also obtained evidence which indicates such an accessory factor closely associated with the I aleurone factor which is linked with waxy, and Coulter (1925) has obtained aberrant segregations in material heterozygous for the C aleurone factor which is also located on the same chromosome.
In plants or animals having free-swimming, mobile sperm, there may be no discrimination on the part of the egg in favor of certain of the male gametes, providing that all are compatible. But all of the sperm are not always capable of reaching the egg. Some are so handicapped as a result of their germinal constitution that, although produced in equal numbers at sporogenesis, they are poorly represented at the time of fertilization.
In the spermatophytes, the gametophyte generation is interposed between the time the gametes are segregated and fertilization, and there is here a relatively large opportunity for the differential elimination of certain classes. In the lower plants the gametophyte generation is of equal or greater importance than the sporophyte generation. In such plants hereditary factors are as important in the development of the one as in the other. Inherited characters which are expressed in the gametophyte generation are described by Wettstein (1924) in the mosses and by Allen (1925) in the liverworts. This generation has become greatly reduced in the spermatophytes, and the hereditary factors are less conspicuous though certainly not entirely eliminated. The pollen tube of the gymnosperms and angiosperms represents the remnant of the gametophyte generation in their phylogenetic ancestors.
Transmissible factors are known to affect all stages of the growing plant. In maize the embryo, the endosperm, the resting stage of the seed, the young seedling, stature, chlorophyll development, and reproductive processes are all controlled by different genes. One would not expect the gametophyte generation, although brief in time and extent of development, to be entirely without differences in its inherited potentialities.
The several Mendelian factors which have been found to have their expression in the gametophyte generation produce a definite physiological effect upon the gametophytes, hastening the growth of the pollen tubes in some plants and retarding it in others.
There is apparently no way in which these factors find expression in the female gametophyte. There is only the one case in Oenothera, previously described, in which the different female gametes are apparently fertilized unequally by pollen of homogeneous constitution and in this case an explanation may be found in differential elimination of gametes, zygotes, or other influences. In the male gametophytes they have no visible effect upon recombination as long as the parental sporophyte is homozygous for these factors. Even when the pollen grains differ, the favorable and unfavorable determiners, in the long run, tend to balance each other so that no gametes would be given a decided advantage.
The markedly greater effectiveness of the plant's own pollen in accomplishing fertilization, which has been noted in experiments with pollen mixtures described in the following chapter, is probably the result of the working of such factors either wholly or in a large part. In practically all of the types tested by applying a mixture of pollen to both lots at least a slight preference was shown for their own pollen, indicating that these gametophyte factors are widely distributed. Abnormal segregations in Oenothera, Melandrium, Datura, rice, barley, and peas afford additional evidence to that obtained from maize. Why they are not a more disturbing element in Mendelian computations is due to the fact that they are seldom closely linked with factors whose segregations can be clearly classified free from other deviating influences. And even when deficient or excessive ratios are obtained, they have been obscured by the prevalent practice of compiling data from many individuals of different pedigree and in successive generations in order to obtain as large numbers as possible. This procedure serves its purpose well. The thoroughgoing Mendelianist seldom fails to obtain "very good ratios." Meanwhile the factors which control the growth of the gametophyte generation in the higher plants have not received the attention they deserve considering the fact that they influence in a measureable degree the way in which inherited characters are recombined. Furthermore, there is the possibility that in these factors we see the beginning of the incompatibility which, when accumulated in sufficient degree, separates different groups by preventing cross-fertilization between them.