Homoeologous Chromosomes From Two Hordeum Species Can Recognize and Associate During Meiosis in Wheat in the Presence of the Ph1 Locus

Abstract

Understanding the system of a basic eukaryotic cellular mechanism like meiosis is of fundamental importance in plant biology. Moreover, it is also of great strategic interest in plant breeding since unzipping the mechanism of chromosome specificity/pairing during meiosis will allow its manipulation to introduce genetic variability from related species into a crop. The success of meiosis in a polyploid like wheat strongly depends on regular pairing of homologous (identical) chromosomes and recombination, processes mainly controlled by the Ph1 locus. This means that pairing and recombination of related chromosomes rarely occur in the presence of this locus, making difficult wheat breeding trough the incorporation of genetic variability from related species. In this work, we show that wild and cultivated barley chromosomes associate in the wheat background even in the presence of the Ph1 locus. We have developed double monosomic wheat lines carrying two chromosomes from two barley species for the same and different homoeology chromosome group, respectively. Genetic in situ hybridization revealed that homoeologous Hordeum chromosomes recognize each other and pair during early meiosis in wheat. However, crossing over does not occur at any time and they remained always as univalents during meiosis metaphase I. Our results suggest that the Ph1 locus does not prevent chromosome recognition and pairing but crossing over between homoeologous. The role of subtelomeres in chromosome recognition is also discussed.

Keywords: barley; chromosome recognition; homoeologous pairing; introgressions; meiosis; wheat.

Figure 1

Hordeum chilense and H. vulgare double monosomic addition lines in wheat and physical location of the HvT01 subtelomeric repeat on barley chromosomes. Hordeum chilense (green) and H. vulgare (red) chromosomes were detected in GISH experiments in somatic chromosome spreads. In addition, the HvT01 subtelomeric sequence from barley was also detected (green). Total genomic DNA was counterstained with DAPI (blue). (a) Double monosomic 7H^ch7H^v addition line, including a diagram showing the HvT01subtelomeric signals in all barley chromosome arms, except in 7H^chL arm. (b) Double monosomic 7H^ch5H^v addition line, including a diagram showing the subtelomeric barley sequence in all barley chromosome arms except in 7H^chL arm. (c) Double monosomic 5H^ch5H^v addition line, including a diagram showing the subtelomeric barley sequence in all barley chromosome arms, except in 5H^chL arm. Bar represents 10 μm.

Figure 2

Behavior of homoeologous and non-homoeologous barley chromosomes during early meiosis in wheat. Hordeum chilense chromosomes are visualized in green and H. vulgare chromosomes are visualized in red. (a) Double monosomic 7H^ch7H^v addition line showing both barley chromosomes un-associated at early pachytene. (b) Homoeologous barley chromosomes are fully associated at late pachytene in the double monosomic 7H^ch7H^v addition line. (c) Double monosomic 7H^ch5H^v addition line showing both barley chromosomes un-associated at early pachytene. (d) Non-homoeologous 7H^ch and 5H^v chromosomes remained un-associated at late pachytene. (e) Double monosomic 5H^ch5H^v addition line showing homoeologous wild and cultivated barley chromosomes half-paired at early pachytene. (f) Double monosomic 5H^ch5H^v addition line showing both homoeologous Hordeum chromosomes fully associated. Bar represents 10 μm.

Figure 3

Detection of the subtelomeric HvT01 sequence in the wild and cultivated barley chromosomes in the 5H^ch5H^v double monosomic addition line. DNA was counterstained with DAPI (blue). (a) GISH of both H. chilense (green) and H. vulgare (red) homoeologous chromosomes initiating pairing during early pachytene. (b) Detection of the subtelomeric HvT01 probe (green) on the same cell. (c) Merge image showing the subtelomeric HvT01 signal (overlay in white) on the terminal region of the unpaired homoeologous wild and cultivated barley chromosome arms (arrowed). Bar represents 10 μm.

Figure 4

Hordeum chilense and H. vulgare chromosome behavior during metaphase I in parental H. chilense and H. vulgare disosomic addition lines in wheat. Hordeum chilense (green) and H. vulgare (red) chromosomes were observed always associated in pairs in all the cells in metaphase I in each H. chilense and H. vulgare disomic addition line. DNA was counterstained with DAPI (blue). (a) H. chilense chromosome 7H^ch disomic addition line. (b) H. vulgare chromosome 7H^v disomic addition line. (c) H. chilense chromosome 5H^ch disomic addition line. (d) H. vulgare chromosome 5H^v disomic addition line. Bar represents 10 μm.

Figure 5

Hordeum chilense and H. vulgare chromosome behavior in double monosomic barley addition lines in wheat during metaphase I. Hordeum chilense (green) and H. vulgare (red) chromosomes remained unassociated in all the cases. DNA was counterstained with DAPI (blue). Centromeres (red) were labeled with RT sequence to show the correct orientation of wheat chromosomes during metaphase I. Subtelomeres on barley chromosomes were labeled in green. (a) Double monosomic 7H^ch7H^v addition line. (b) Double monosomic 7H^ch5H^v addition line. (c) Double monosomic 5H^ch5H^v addition line. Bar represents 10 μm.

Figure 6

Behavior of Hordeum chilense and H. vulgare chromosomes in double monosomic barley addition lines in wheat during anaphase I of meiosis. Examples of barley chromosome segregation after metaphase I. Hordeum chilense and H. vulgare were visualized in green and red, respectively. DNA was counterstained with DAPI (blue). (a) Double monosomic 5H^ch5H^v addition line, (b) Double monosomic 7H^ch7H^v addition line, (c) Double monosomic 7H^ch7H^v addition line; (d) Double monosomic 7H^ch7H^v addition line. (e) Double monosomic 7H^ch5H^v addition line. (f) Both 7H^ch5H^v barley chromosomes remained delayed. (g) Both 7H^ch7H^v barley chromosomes remained delayed and a misdivision of chromosome 7H^v was also observed. One or both barley micronuclei were positioned in the equatorial region on telophase I in (h) Double monosomic 7H^ch5H^v addition line, (i) Double monosomic 7H^ch7H^v addition line, and (j) Double monosomic 7H^ch7H^v addition line. The subtelomeric HvT01 probe was used in GISH experiments performed in cells in telophase I from the double monosomic 7H^ch5H^v addition line to visualize (k) 5H^v chromosome misdivision or (l) 5H^v chromosome segregation. Bar represents 10 μm.

Figure 7

Behavior of H. chilense and H. vulgare chromosomes in double monosomic Hordeum addition lines in wheat after the second meiosis division. Examples of the most frequent observations for both H. chilense (green) and H. vulgare (red) chromosomes are shown. DNA was counterstained with DAPI (blue). (a) Hordeum vulgare and H. chilense chromatin detected in four and two PMCs, respectively, in the double monosomic 7H^ch7H^v wheat line. (b) Hordeum vulgare and H. chilense chromatin detected in four and in two PMCs, respectively, in the double monosomic 5H^ch5H^v wheat line. (c) Four H. chilense signals and two H. vulgare signals were observed in the double monosomic 5H^ch5H^v wheat line at the same frequency as reported in b. (d) Wild and cultivated barley signals are present in the same nucleus and only one each in another two different nucleus. Bar represents 10 μm.