Cucurbit Genetics Cooperative Report 19:73-77 (article 27) 1996
Oved Shifriss
21 Walter Avenue, Highland Park, NJ 08904
The B genes are nuclear mutants, and each B gene conditions precocious depletion of chlorophyll. In fruits, these genes can be expressed in all known genetic backgrounds. But in some backgrounds they can also be expressed in one of more other plant organs. Historically, symbol B represented a genetic potentiality for bicolor fruits in which the non-green portion is depleted of chlorophyll.
Two unlinked B genes were identified, B1 and B2. B1 originated in C. pepo L. and B2, in C. maxima Duch. ex Lam. Gene B2 is not an uncommon mutation in open-pollinated populations of some standard B2+ cultivars. Therefore, one may conclude that such a mutation was the origin of all our B2 cultivars. The literature hints to the past existence of a few edible B1 cultivars in C. pepo, but no clue is given as to their origin. Nonetheless, it is evident from genetic data that the presently utilized B1 had a two-step origin in the inedible ornamental gourds of C. pepo var. ovifera (L.) Alef. The first step was a mutation from B1+ to B1w, the later being a gene for a weak expression of chlorophyll depletion and bicolor fruits. The second step was a mutation from B1w to B1, a gene for stronger expression and uniformly pigmented yellow fruits. There is also evidence indicating that the B1 locus is unstable in some backgrounds and stable in others. B1w and B1 were transferred to more than ten edible cultivars of C. pepo. In addition, B1 and B2 were transferred to several cultivars of C. moschata Duch. ex Poir,, a species that has not been reported to generate B mutations. Recent field observations suggested that ‘Gold Striped Cushaw’ of C. argyrosperma Huber carries a weak B gene whose phenotype resembles that of B2. The seed of this cultivar was obtained from Glenn Downs.
Potentially, each B gene brings about a profound change in the ontogenetic timing of chlorophyll depletion during fruit development. As a result, three distinct developmental phases of fruit pigmentation are recognized in cucurbita: the standard phase, the precocious phase #1 and the precocious phase #2 (Figure 1).
The standard phase of pigmentation represents the post-anthesis phase of fruit development in all B+B1+B2+N2+(B+) cultivars. Prior to anthesis the ovaries of these cultivars are consistently green. From anthesis and on, the fruits are either persistently green or turn to other colors (white, tan, yellow or golden) depending upon the presence of specific pigment-controlling genes such as w and y for persistent green and W and Y for white or yellow colors. Whether the change in color occurs at anthesis or on approaching maturity the fruits are uniformly pigmented. Furthermore, the change in color and chlorophyll depletion appears to occur simultaneously.
The precocious phase #1 and the precocious phase #2 represent cultivars whose genotypes are B1B1 B2+N2+(B1) and B1+B1+ B2B2 (B2) respectively. Chronologically, phase #1 ends a few days prior to anthesis and phase #2, a few days following anthesis. Within these phases, B1 and B2 can be expressed at different times: the later the expression the lesser is the extent of yellow pigmentation of the ovary. Consequently, B1 and B2 plants can differentiate either uniformly yellow ovaries or bicolor (green-yellow) ovaries. As these ovaries grow through and beyond anthesis, the color of their maturing fruits is affected by the pigment-controlling genes that are active at this stage of development. For example, the ovaries of Atlantic Giant, B2B2, are bicolor whereas the mature fruits are uniformly white. If the B genes fail to “turn-on” during their limited phases for potential action, the color of the B fruits would be indistinguishable developmentally from that of B+ fruits.
The ontogenetic timing of B expression is governed by several factors: the strength of the B allele, the dosage of B, a group of nuclear timers (regulators) and the environment. The nuclear timers were formerly known as modifier genes that affect the extent of precocious pigmentation over the surface of the fruit: the Ep and ep modifiers of B expression. The heterozygote B1wB1+ of some ornamental gourds is particularly prone to delay the expression of B1w and is apt to exhibit a remarkable phenotypic plasticity, bearing variable bicolor fruits as well as green fruits in an unpredictable order. The homozygote B2B2 of PI 165558 is associated with one of the earliest known expression of B2 and is highly stable phenotypicaly, bearing uniformly golden fruits exclusively.
White bicolor fruits vary greatly in extent of green and non-green (yellow or golden) areas, the distribution pattern of these areas is subject to distal-proximal polarity, green in the lower or distal portion of the fruit and non-green in the upper region, towards the proximal end. The bicolor pattern is further affected by other factors including fruit shape, e.g., the “ring” phenomenon is limited to long-necked (ladeled) fruits as in ‘Spoon’, In a rare case, the common polarity radically moves upward: the fruit being essentially green and the peduncle, uniformly golden (Figure 2). In another rare case, the common polarity is completely reversed, golden in the distal portion and green in the upper region of the fruit.
Interactions between the B genes and other genes lead to numerous phenotypic effects most of which are horticulturally “bad” but some are “good”. The basic biological mechanisms controlling these manifold effects are not understood. Yet, pragmatically, one can be guided by the results of breeding experiments. These results imply that the cucurbita genome is highly resourceful and that through genetic recombination the “bad” effects can often be eliminated while the “good” effects are sustained or enhanced.
The ovaries of many BB homozygotes develop poorly and the size of their fruits is smaller than that of comparable B+B+ fruits.But the ovaries of ‘Atlantic Giant’, B2B2, develop normally and their fruits are the largest on this planet.
In B+ cultivars of Winter Squash, young fruits are low and mature fruits are high in levels of flesh carotenoids. In B+ cultivars of Summer Squash, young and mature fruits are low in flesh carotenoids. The external fruit color in these two groups of cultivars may be green, tan, yellow, or golden. The genetic mechanism that controls the level of flesh carotenoids in B+ cultivars is not known. A new system for high levels of flesh carotenoids was developed through a particular interaction between the B genes and the L genes. In this B-L system, both young and mature fruits contain high levels of flesh carotenoids and their external color is persistently golden. The L genes (L-1 and L-2, Paris) condition a high rate of pigment accumulation during fruit development. And the best sources of L genes are cultivars whose fruits are dark green, gradually becoming solid black at maturity.
The yellow and golden fruits of all presently known B+ cultivars and most B cultivars are subject to green discoloration due to virus infection. But fruits of some BB liens of C. pepo and C. maxima are resistant in varying degrees, Although resistant lines are usually homozygotes, BB, not all homozygotes are resistant. The decisive factor appears to be the ontogenetic timing of B expression: the earlier the expression of B the higher is the probability for resistance. From this perspective, the operative system is the interaction between the B genes, their nuclear timers. and the environment. Field records showed that the fruits of PI 16558, B2B2, possess the highest known level of resistance to virus-induced greening. Do the B genes carry virus information?
Some interactions between the B genes and other genes appear to enhance female expression. Other interactions appear to affect fruit quality in diverse ways. But the evidence for these interactions has not been critically examined.
The environment plays an important role in the expression and regulation of the B genes. Among potential influences, the effects of temperature have been demonstrated in growth-chamber studies and confirmed ny field observations. Low temperatures (e,g,, 12 C), high temperatures (e,g,, 35 C) and intermediate temperatures can have different effects on the expression of the B genes, depending on the genetic background. Light, may also be an important influencing factor but I am not aware of any experimental data on the effects of light on the expression of the B genes.
Apart for some exceptions, the exposure of B cultivars, BB or BB+, to a progressive increase of temperature, from low to high, is associated with a corresponding decrease in extent of precocious fruit pigmentation.
When young plants of B1+B1+ andB1B1 inbreds are exposed to low temperatures, the leaf-blades of the two groups of inbreds exhibit yellow or golden spots. But the incidence of spots is 3-to 10-fold higher in B1B1 than in B1+B1+ leaves. No such spots occur in B1+B1+ and B1B1 leaves at high temperatures. Are the induced spots in B1+B1+ leaves responses to “residual heredity” that is genetically related to B1?
When young plants of genetically diverse B1B1inbreds are exposed to low temperatures, the leaf-blades of some inbreds are yellow whereas the leaves of other inbreds are green. The yellowing is expressed either as diffusion affecting the entire leaf surface or as variable patterns, often as “vein designs.” No such yellowing occurs at high temperatures. A study of inheritance, based on one relevant cross, showed that these alternative responses are conditioned by a pair of nuclear genes. The recessive gene behaves as a selective suppressor. This monog4nic inheritance is probably an over-simplification because temperature-sensitive inbreds differ in degree of their sensitivity. In sensitive inbreds, both spotting and yellowing occur in different leaves of the same plant.
Selective regulation of B1 expression is limited to leaf-blades. However, there exist rare cases in which stems and petioles are yellow. One of these cases originated as mutation in a green-stemmed, predominantly-female (PF) B1B1 inbred. Like its parent, the mutant inbred is PF and B1B1. But unlike its parent, the mutant stems and petioles gradually turn yellow in proximity to well-developed pistillate flowers and fruits (Figure 3; some of the lower pistillate flowers were removed in order to expose the main stem). According to present interpretation: (a) precocious chlorophyll depletion is preceded by a diffusible substance; (b) the mutant produces a larger amount of this substance than its parent; (c) the substance is diffused from pistillate flowers to adjacent regions; and (d) a high concentration of pistillate flowers promotes the flow.
Selective regulation of B2 expression can affect leaf-blades, petioles, stems and staminate flower buds, But the precise regulatory mechanism is not yet understood.
The American B2B2 cultivars such as ‘Boston Marrow’, ‘Golden Delicious’, ‘Golden Nugget’ and ‘Pink Banana’ produce green stems and green leaves in diverse environments of low, intermediate and high temperatures. By contrast, PI 165558, B2B2, a cultivar from Almora, India, is more sensitive to temperature fluctuations. When this cultivar is grown in an environment of intermediate temperatures, it produces precociously yellow stems and green leaves. If, in the same environment, this cultivar is exposed to a few cold nights (2-3) of low temperatures, is subsequently continues to produce yellow stems but it also produces, for a short period, some yellow-spotted leaves as well as some completely yellow leaves. But when this cultivar is grown in an environment of high temperatures, it produces green stems and green leaves. Unlike the yellowing of stems in the B1B1 mutant described above, stem yellowing in PI 165558 occurs during both the vegetative and reproductive stages of plant development, and it is clearly not associated functionally with pistillate flowers.
When some yellow-stemmed B2B2 inbreds – derivatives of crosses between B2+B2+ cultivars and PI-165558, B2B2 – are grown under field conditions of intermediate temperatures, all their leaves gradually turn yellow. These inbreds can be reproduced by seed, but with great difficulty. When the same inbreds are grown in an environment of low temperatures, their populations consist entirely of lethal seedlings.
The B2 of PI 165558 was transferred to the B2+B2+ cultivar of ‘Green Delicious’ (Munger’s strain). The new precocious B2B2 inbred of ‘Green Delicious’ background – PGRD – produces green stems and green leaves in environments of diverse temperature variations as do other B2B2 cultivars, which are essentially bicolor-fruited, the PGRD inbred produces uniformly golden fruits. Is the B2 allele of PI 165558 stronger than that present in most other B2 cultivars?
The transfer of B2 to C. moschata revealed two facts. First, C. moschata is an exceptionally rich source of genetic elements that selectively activate the expression of B2 in stems. Second, some of these genetic elements impart great phenotypic stability to the expression of B2 in stems even at high temperatures. If we consider both C. maxima and C. moschata, we see a continuous series of variation in the expressivity of B2 in stems, from zero expressivity to 100% expressivity in diverse environments. All intermediate grades are environmentally sensitive in different degrees. The simplest hypothesis is that the expression of B2 in stems is governed by numerous genetic regulators that interact with the environment.
In 1983, a mutant was found in an F2 of ‘Bicolor Spoon’, B1wB1w , x ‘Table King’, B1+B1+. This mutant was named ‘Golden Crown’ (Figure 4): it differentiates a number of strictly green leaves along the main stem followed by a transitional phase towards completely yellow or golden leaves at the top (the golden crown). Phenotypicaly, this mutant resembles many ornamental cultivars including Amaranth us tricolor var. spendens and the so-called “ornamental cabbage” or “ornamental kale” cultivars (Brassica oleracea, acephala group). One can easily obtain a true-breeding line of ‘Golden Crown’. But serious difficulties were encountered for a long time in attempts to decipher the genetic basis for this mutant. With some temerity one can now claim that ‘Golden Crown’ is conditioned by a single nuclear gene.
Recently, a cross was made between ‘Fordhook Zucchini’ (Supernak’s strain) as seed parent and ‘Golden Crown’. The F1 plants produced green leaves exclusively. Among 95 F2 plants, not a single ‘Golden Crown’ individual was found, but one bicolor-fruited plant was clearly identified. If these results can be confirmed, (a) did ‘Golden Crown’ originate as transposition of some element related to or part of B1w ?, and (b) did the bicolor-fruited plant in the above F2 originate as reverse transposition?
Figure 1. Illustrating three phases of fruit development in Cucurbita. Fruit pigmentation other than green can occur during either one of these phases depending on the genotype and the environment.
Figure 2. Illustrating a fruit in which precocious depletion of chlorophyll largely affects the peduncle. Fruits of this pattern were found in a rare segregate obtained from the interspecific cross: C. maxima (“Pink Banana”) x C. moschata (‘Chirimen”).
Figure 3. Illustrating complete association between precocious depletion of chlorophyll in stems and predominantly- female expression. This association exists in a rare inbred of C. pepo.
Figure 4. Illustrating the ‘Golden Crown’ mutant of C. pepo.
Acknowledgement
I thank Herbert H, Bryan and Waldemar Klassen for enabling me to grow my squash germplasm for two seasons, from 1994 to 1995, at the Tropical Research and Education Center of the University of Florida in Homestead.