Spm and I element changes with the a-m2 8004 allele in maize

Summary Among the mobile element systems in maize, the En (Spm) system (En — the regulatory element and I the receptive element — a nonfunctional En) has several interesting aspects of control of gene expression (En and Spm are homologous in structure and activity). One of the alleles arising from the Spm group included the a-m2 8004 allele that has a low spotting pattern and unique ringed areas. The interest in this allele is that Spm or En will induce it to co-express the A phenotype and mutability. Several exceptions of the allele were analyzed. Two are Spm changes and two are I changes. The analysis shows that the heritable changes include I changes that are co-expressed in various grades of color and different degrees of mutability. All these changes occur with I at the locus. The Spm changes also include changes in mutability patterns and a mottling pattern.Journal Paper No., J-11792 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa, Project No. 2381     

Enhancer transposable element induced changes at the A locus in maize: The a-m-1 6078 allele

Summary Transposable (controlling) elements in maize are highly variable in their versatility in inducing changes at any locus. With the Enhancer (En) (Suppressor-Mutator-Spm) controlling-element system,a multitude of changes result from the action of En on specific alleles. One such allele, McClintock's a-m-1 6078 allele, illustrates the diversity of events that can take place that involve three parameters; namely, spotting-timing, spotting-frequency, and background pigment formation with and without En. A series of derivative alleles has been isolated and described according to the three paramters. The multitude of changes that has been isolated illustrates the striking versatility of these transposing elements to initiate changes at the controlled locus.Journal Paper No. J-11135 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa, Project No. 2381     

Chromosome labeling with transposable elements in maize

Transposable elements randomly insert into a targeted locus at a frequency of 10^-6 to 10^-5. The En element has been shown in previous studies to transpose more frequently into closely linked sites. Thus, it is appropriate to place an En element onto each of the 20 chromosome arms in maize to maximize tagging efficiency. This is called chromosome labeling for tagging purposes with transposons. After a chromosome arm has been labeled with a transposon, genes residing in that arm will have a greater chance to be tagged by the transposon. To date, all of the maize chromosome arms have been labeled with at least one of five Encontaining alleles. The elements were linked to the arms using reciprocal translocations. The usage of these arm-labeled lines is discussed in the context of gene tagging.Journal Paper No. 15224 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa; Project No. 3176     

Transposition of the enhancer controlling element system in maize

Summary Controlling elements have the ability to activate or inactivate standard genes. One of the unique features of controlling elements is their ability to transpose from one position to another on the same chromosome or to another chromosome. In this study, the transposition of the controlling element En was examined in its transposition from its initial site at the al locus to a primary site and then to secondary sites, all within the distal two-thirds of the 3L maizechromosome arm.Stable germinal mutations representing losses of En were selected from three different autonomously mutating al alleles. The insertion of En at a new location was confirmed by crosses of the stable mutants to responsive tester lines. The position of En at the primary transposition site was determined by backcrossing to an al et line using standard three-point tester crosses. Secondary transpositions were detected by Chi-square comparisons of progeny to parental En linkage values.Although En positions were found throughout the segment of chromosome examined, their distribution was not random and some regions of the chromosome were more likely to contain an inserted En. Ens from all three autonomously mutating source alleles showed the same regional preferences. Distributions of transposed Ens on chromosome 3L from the three original unstable al alleles were not significantly different in Chi-square tests. Both primary and secondary sites of insertion were located within these regions. No differences were found between the distribution of primary sites of insertion and the distribution of secondary sites. Statistically significant differences were found between the distributions of transposing and non-transposing Ens.Formerly graduate student, ISU, now with Monsanto Agricultural Products Co., 800 N. Lindbergh Blvd. St. Louis, MO 63166, and Professor of the Department of Agronomy, Iowa State University, Ames, 1A 50011     

The inveterate wanderer: study of Enhancer wandering on chromosome 3 in maize

The transposition of the maize transposable element Enhancer (En) had been focused on one chromosome 3 for several generations. From the a1-m(Au) allele with an autonomous En, a new En reporter allele a1-m(r)3927-1, was isolated that undergoes very infrequent and late excision events, producing one or two small spots in the aleurone. This allele is seriously impaired in its capacity to excise. Coincident with the origin of this allele, an En was located at a site close to the a1 locus. From this initial insertion site, the movement of this En was followed for three to four generations in 974 families with a higher transposition rate of this En (50% of the testcross progeny) than that found in a previous study of En transposition. This is the first case reported where a particular En was followed for more than three generations. The higher rate of wanderings of this En along the same chromosome led to the term vagabond En (En ^vag ). Genetic evidence that En may transpose from a replicated donor site to an unreplicated site is provided. Speculative mechanisms on the origin of a1-m(r)3927-1 and En ^vag are discussed.Journal Paper No. J-15864 of Iowa Agricultural and Iowa Economical Experiment Station Project #3176     

Change in State following Transposition of a Regulatory Element of the Enhancer System in Maize

From an original A2 allele (colored aleurone), a mutable allele, a2-m-4 1629, that changes from a2 to A2 is described. Mutability is expressed as a very distinct pattern limited to the last cell division.—The mutability of a2-m-4 1629 is autonomously controlled by an En at the a2 locus. This En, inactive on standard a testers for En, is partially active on a2-m-1, an a2 tester for En, and expresses varied levels of activity from limited to nearly full suppression of the a2-m-1 color phenotype.—When the En of the a2-m-4 1629 allele transposes from the a2 locus, it behaves, at the new position, like a standard En in triggering a2-m-1, a-m-1 and a-m(r), which express colored spots on a colorless background. The activity of En is therefore different following the change in chromosome location. This finding supports the "position" hypothesis that has been proposed to explain diverse patterns observed among controlling elements. In this case mutation is related to the terminal cell state and not to tissue differences as shown with some phase-variation regulatory elements.     

The Fcu controlling-element system in maize

Summary The relationship between the Fcu controlling-element system and the spotted-dilute R system was investigated. The Fcu controlling-element system consists of the receptor element allele r-cu and the regulatory element Fcu. The equivalent components of the spotted-dilute R system are respectively R-r#2 (or R-r#2 Dil) and Spf. The R-r#2 allele of the latter system was shown to be responsive (mutable) to Fcu, provided that it has had an uninterrupted association with Dil or Dpf as evidenced by the color variegation of the aleurone tissue. The reverse test, in which the r-cu allele of the Fcu controlling element system was tested for its response to Spf, proved negative. This was surprising in view of the relationship and specificity between systems. The possibility was considered that maize controlling elements may have different sizes as is known for bacterial insertion sequences. —The variable dilute pigmenting capacity of the r-cu allele also was studied. A given level of r-cu-induced pigmentation, despite the wide range in pigmentation expression, was found to be generally non-heritable, as based on a test of correspondence between parent and progeny.Journal Paper No. J-9204 of the Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa. Project No. 1884Now with Funk's Seeds, Casalmorano (Cremona), Italy     

Genetic regulation of somatic mutability of two Mu-induced a1 mutants of maize

Summary Previous studies of stocks of two Mutator-induced mutable a1 alleles (a1-Mum2 and al-Mum3) gave results consistent with the presence of one or more autonomous elements regulating the expression of mutability. This article reports on the results of studies designed to map these autonomous elements by using a series of waxy marked translocations. Linkage of waxy with autonomous elements was found for a1-Mum2 by using the translocations wx T2-9d, wx T4-9e and wx T4-9b. Several different linkage values were found in crosses involving wx T2-9d, suggesting that autonomous elements have transposed to different locations on chromosome 2. Linkage of autonomous elements with waxy was found for a1-Mum3 using translocation wx T2-9d. Again, several different linkage values were found. Some of these values were the same as those observed for a1-Mum2, but some were unique. In some crosses, the number of autonomous elements increased by one or two unlinked elements in addition to the linked element in one generation (i. e. the generation of the cross to the translocation series). Such an increase in number is probably the result of transposition of the original autonomous element to an independent locus while retaining the autonomous element at the original locus. Reduction in the number of autonomous elements is probably the result of the independent assortment in crosses of plants with two or more autonomous elements.Journal Paper No. J-14569 of The Iowa Agriculture and Home Economics Experiment Station, Ames, Iowa Project No 2870     

Analysis of a case of somatic instability in a strain of maize from India

A case of somatic instability affecting aleurone colour in a strain of maize from India with flint background was analysed. The somatic instability is localized to theC ¹ (Inhibitor) allele ofC locus on the short arm of chromosome 9. Molecular tests indicated thatAc is not present in the Indian stock and the evidence is consistent with the involvement of theEn (Spm) transposable element in the instability. The presence of theEn (Spm)-like element in the stock would suggest that these elements have been present in the maize genome for a long time. A new allele ofshrunken (sh1) gene with a somewhat unorthodox breeding behaviour is also described.     

Somatic Mutator activity expression is dependent on the strength of Cy trans-active signals in maize

Summary Two receptor element alleles (vp-rcy and bz-rcy) that respond to the trans-active element (Cy) controlling Mutator activity were used to analyze the strength of trans-active signals from Cy elements derived from a Mutator active line. Evidence is presented that the Mutator population tested consists mainly of a class of weak Cy elements designated as Cy:Mu. When Cy:Mu element are present in only a few copies, the strength of the combined transposition signal is weak. It is only when these active elements have a high copy number that the overall transposition signal is sufficiently strong enough to elicit a high frequency of transposition events. This study seeks to investigate the nature of the trans-active signal from Cy:Mu elements. The implication of these results for molecular studies is discussed.Journal Paper No. J-13083 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA, Project No. 2381     

Genetic and molecular analysis of a three-component transposable-element system in maize

Summary Two different factors control the mutability of an unstable allele (c2-m8810581) of the C2 gene of maize. Both an autonomous En/Spm element and an unrelated independent factor, named Mediator, are coordinately required for the excision of the insert in c2-m881058Y. According to genetic analysis, Mediator does not have the suppressor (S) function or mutator (M) function of En/Spm. Mediator has no effect on the timing or frequency of excision of Enl, En-low, or various I/dSpm elements. Hence, Mediator only mediates a specific interaction between En and the insert at c2m881058Y. Molecular analysis of c2-m881058Y has revealed a 3.3 kb, complex, En-related receptor element inserted into the second exon of the C2 gene. The ends of this element are homologous to the ends of En/Spm, but an internal l.7 kb region shows no En/Spm homology. A great degree (11–14%) of nucleotide changes, relative to Enl, occur within and between the 12 bp TNPA binding motifs. Alterations of these critical cis-determinants may account for the need for a helper factor for excision. This element is named Irma, for Inhibitor that requires Mediator also, and represents a unique, low copy number class of receptor element.     

Differential excision patterns of the <Emphasis Type="Italic">En</Emphasis>-transposable element at the <Emphasis Type="Italic">A2</Emphasis> locus in maize relate to the insertion site

Defined mutant alleles with resident transposons display characteristic patterns of germinal and somatic reversion, and heritable changes in the timing and frequency of reversions, which have been termed “change of state” by McClintock, constantly arise. Several mechanisms were proposed to account for these changes. They may be ascribed to the structure and composition of the elements themselves (composition hypothesis) or to their location (position hypothesis). In the current study, insertion positions were determined for three autonomous En-controlled mutable alleles of the A2 locus in maize that show different somatic reversion patterns. A relationship was observed between En insertion positions in the single coding region of the intronless A2 gene and anthocyanin variegation patterns in the aleurone. An insertion in the 5′ region of the coding sequence produced a very late somatic variegation pattern, whereas two early variegation patterns were caused by En insertions in the 3′ region of the coding sequence.     

Controlling elements in a tribal maize from Colombia: fcu , a two-unit system

The genetic basis for a striking aleurone color variegation of a maize population from Colombia is described. The variegation is the result of the interaction of a particular r allele, designated r-cu , and a specific factor, Fcu (Factor Cuna), that collectively compose a controlling element system. In the absence of Fcu, a variable dilute (nonvariegated) pigmentation is expressed in the aleurone. The degree of variegation is determined by differences in the dosage of the Fcu element; differences in variegation between kernels with one r-cu vs. two r-cu's cannot be detected. Because of the specificity of the interaction, Fcu is different from the three other well-known controlling elements, Dt, Ac and En(=Spm), and therefore represents a fourth controlling element system in maize. In addition to Fcu, the Cuna tribal maize contains two other controlling elements, Dt and a very weak En. The prevalence of controlling elements in natural maize populations is discussed.     

An altered state of a specific en regulatory element induced in a maize tiller

There are numerous states of the regulatory element, Enhancer (En). With specific receptor alleles, such as a2m(r-pa-pu) or a2m(r), specific mutability patterns are expressed. One specific derivative En allele, En-v (En-variable), was originally identified with a coarse pattern of mutability with the a2m(r-pa-pu) allele and giving progeny with varied En expression (standard to reduced within an ear progeny). Derivatives of En-v were subsequently found on numerous occasions to give only a very reduced expression (fewer mutant spots) with the a2m(r-pa-pu) allele in the ears derived from the main stalk of the corn plant. When a comparison is made of the effect of this changed En-v state between tiller ears and main stalk ears of the same plant, the tiller ears show an increased level of En-v expression (coarse pattern), while the main-stalk ears continue to show the very reduced level of En-v expression (low frequency of very late variegation). This increased level of mutability of the tiller ears is maintained when transmitted through the main-stalk ear in the subsequent generation. These results indicate that heritable alterations of controlling elements can be produced by endogenous environmental factors present during normal plant development.     

Genetic control of immunoglobulin allotypes in the fowl

Eight immunoglobulin allotypic specificities have been identified in the fowl by isoimmunization. Aa1 and Aa2 are controlled as codominants at the a locus, Ab1 and Ab2 at the b locus, and Ac1, Ac2, Ac3, and Ac4 at the c locus. Column chromatography and ultracentrifugation indicate that the specificities at the a locus are located on molecules corresponding to IgG with sedimentation coefficients 7 S. Immunoelectrophoresis results also indicate that we are dealing with an immunoglobulin G molecule. Further tests are underway to resolve this beyond doubt.Journal Paper No. 5405 of the Iowa Agricultural and Home Economics Experiment Station, Ames, Iowa. Project 1039. Supported in part by National Science Foundation Grants GB 318 and GB 4450.     

The a2m(r-Pa-Pu) Allele of the En-Controlling Element System in Maize

The Enhancer (En) controlling element system in maize includes numerous alleles, one of which, a2m(r-pa-pu), was recovered as a derivative of the a2m(1 1511) allele. The behavior and composition of this allele is described. In the presence of the independently inherited regulatory element, En, the a2m(r-pa-pu) allele mutates and expresses in the aleurone purple, pale and colorless sectors within a colorless background. In the absence of En, the pigmentation is uniformly pale. Colored and colorless germinal mutations of the a2m(r-pa-pu) allele were recovered, although the derivative alleles tested did not respond to En. Uniformly colored derivatives, designated A2', arise only from states of a2m(r-pa-pu) that respond to an active En at relatively early stages in plant development, suggesting that somatic and germinal mutations may arise through similar events. Mutations to A2-expressing alleles appear to occur relatively late in plant development, that is, at or near meiosis, since recognizable mutant sectors do not appear on testcross ears. Kernels exhibiting such mutants are randomly distributed over the ear. The resulting A2' alleles express pigmentation ranging in a continual sequential series from dark pale to intense purple.     

Occurrence and behavior of the components of the 02-m(r)-Bg system of maize controlling elements

Summary o2-m(r) is an unstable allele of the O2 locus responding to the regulatory element Bg by somatic reversion. The spontaneous occurrence and the properties of the components of this system of controlling elements have been investigated. The system appears to have some degree of specificity for the O2 locus, because the majority of spontaneous O2 mutations are responsive to Bg. The component at the controlled locus undergoes frequent changes in state, while the Bg element appears more stable. Bg activity was revealed in 11 out of 108 open-pollinated varieties of maize. Most of the newly isolated Bg elements are linked with the O2 locus. The timing of induction of reversion events (which are restricted to mitotic division leading to egg or pollen maturation and to the developing endosperm) appears to correlate with the degree of linkage between Bg and the O2 locus. Germinal reversions of o2-m(r) to wild type give rise with a frequency around 5×10^-4 to unstable phenotypes. Some peculiar features of the o2-m(r)-Bg system of controlling elements are discussed.     

Genetic instability at the opaque-2 locus of maize

Summary A case of genetic variegation discovered at the opaque-2 locus of maize that includes a two-element system with a receptor and regulatory element is described. The somatic mutability depends on the existence of two genetic factors: a responsive allele (with receptor element), o2m(r), and a regulatory element, Bg, that induces mutability of o2m(r). In the absence of Bg, o2m(r) is indistiguishable from the recessive alleles of the O2 locus; in the presence of the regulatory element, o2m(r) mutates giving rise to sectors of flint-like endosperm in an opaque back-ground. The regulatory element Bg may be located independently or at the controlled locus. The genetic properties of the new system, somatic mutability, transposition, existence of different patterns of mutability, are apparently similar to those previously described in maize for the classical systems of controlling elements. In addition, the recovery of the o2 mutability from crosses between spontaneous o2 alleles suggests that transposable genetic elements may be involved in the origin of natural mutability.     

Germinal derivatives of the En controlling-element system in maize: Characterization of colored,pale and colorless derivatives a2-m

Summary Several stable germinal derivatives, with varied phenotypic expression have been recovered from testcrosses of four different a2 mutable alleles (a2m-7 8018, a2m-6 8144, a2m-6 8140, and a2m-1 1511) under the control of the En controlling-element system. These are colored, pale, and colorless aleurone types; the former two represent mutations of a2-mA2 type. The phenotypic variation among colored and pale derivatives has been confirmed by quantitative determination of anthocyanin content in the aleurone tissue of the kernels.The data suggest that significant variation exists between phenotypically similar colored derivatives arising from different mutable alleles as well as from the same source allele. Every tested colored derivative is significantly different from the pale, colorless, and colored control (W22 color converted line). Genetic analysis of these colored derivatives suggests that all of them contain one or more En and, hence, they are characterized as permanent changes of a nonresponsive (nr) type. Their frequencies are variable among the testcrosses.Similarly, the pale derivatives, which are clearly different from colored and colorless, also show significant differences in terms of anthocyanin content. Significant differences exist between pales arising from the same allele as well as between pales arising from different alleles. All the tested pales are significantly different from colorless and colored derivatives. Thus, the pales, as well as colored derivatives, represent a differential impairment of anthocyanin synthesis in the aleurone tissue. The genetic analysis confirmed that all the pales, except the pales of a2m-1 1511, lack the capacity to respond to En. These pale derivatives can be with or without En.Qualitative differences among the colored, pale, and colorless types have been investigated by thin-layer chromatographic and spectroscopic techniques. The results suggest that there are no qualitative differences, in terms of anthocyanin pigments, between colored, pale, and colored control. All accumulate the same anthocyanins (namely, cyanidin-3-glucoside and pelargonidin-3-glucoside in appropriate proportions depending upon the Pr and pr constitution).Journal Paper No. J-8431 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA 50011. Project No. 1884: Support from NSF GB38328.     

Rearrangements at a hobo element inserted into the first intron of the singed gene in the unstable sn 49 system of Drosophila melanogaster

The cytological structure of the X chromosome and the DNA organisation of the singed locus were examined in five singed bristle mutants of Drosophila melanogaster. These mutants are all derived from the unstable mutant singed-49, isolated from a wild population in the Russian Far East in 1975. Rearrangements were found at a site within the first intron of the singed gene, where a hobo element is inserted in these mutants. One rearrangement, which is associated with a strong bristle phenotype, has an inversion between 2D and the location of singed at 7D, which separates the singed promoter from the singed coding region. Two phenotypically wild-type derivatives have smaller rearrangements within the first intron which do not appear to interfere with singed expression. Two derivatives with bristle phenotypes have more complex rearrangements, and one of them shows a dominant or antimorphic phenotype. DNA blotting and in situ hybridisation experiments show that, in addition to these rearrangements at a hobo element inserted at singed, other hobo elements in these strains have been mobilised. This system is therefore similar to others in which functional hobo elements continue to transpose, resulting in elevated rates of mutation and chromosome rearrangement. Received: 19 February 1997 / Accepted: 8 October 1997