Chromosoma (1990) 99:365-370
CHROMOSOMA © Springer-Verlag 1990
Synapsis in single and double heterozygotes for partially overlapping inversions in chromosome 1 of the house mouse Pavel M. Borodin, Ivan P. Gorlov, and Tatyana Yu. Ladygina Institute of Cytology and Genetics, USSR Academy of Sciences, Siberian Department, Novosibirsk 630090, USSR ReceivedJanuary 16, 1990/in revised form March 4, 1990 Accepted April 11, 1990 by H.C. Macgregor
Abstract. Electron microscopic (EM) analysis of synaptonemal complexes (SC) in single and double heterozygotes for the partially overlapping inversions In0)lIcg, In(l)lRk and In(1)12Rk in chromosome 1 of the house mouse reveals that synapsis and synaptic adjustment are dependent on the size and location of the inversions and interaction between the latter. In(1)lIcg contains insertions of the inverted repeats Is(HSR; 1C 5) lIcg and Is(HSR;1D)2Icg and an inverted euchromatic region. Synaptic adjustment of the D-loops by shortening of the asynapsed segments of the lateral elements belonging to the insertions occurs at the late zytogene to early pachytene stage. Synaptic adjustment of the inversion loops takes place at early to late pachytene. A delay in adjustment was found in the double heterozygotes ln(1)lIcg/In(1)lRk and In(1)lIcg/In(1)12Rk. A correspondence between the lifespan of asynapsis in inverted regions and the probability of association of XY and heteromorphic bivalents was revealed. Introduction Using a surface-spreading technique, loop formation and its resolution at the pachytene stage in male mice heterozygous for the inversions In(1)lRk and In(2)5Rk (Poorman et al. 1981a; Moses et al. 1982) and tandem duplication in chromosome 7 (Poorman et al. 1981b; Moses and Poorman 1981) has been described. Similar observations have been reported in heterozygotes of an inverted duplication of a chromosome 1 homogeneously staining region (HSR) (Borodin et al. 1990). The process of loop resolution connected with partial desynapsis and followed by non-homologous resynapsis has been called "synaptic adjustment" (Moses et al. 1982). It has been suggested that synaptic adjustment is a general phenomenon among different species (Moses 1980). However further studies in inversion heterozygotes have revealed that the fate of the loop during meiotic prophase may vary depending on the species and the type of inversion. Offprint requests to: P.M. Borodin
Anderson et al. (1988) distinguish four types of synaptic behaviour of inversion loops: (1) homosynapsis replaced by heterosynapsis. In addition to the above studies this kind of synaptic adjustment is displayed in mice (Chandley 1982; Tease and Fisher 1986), human (Gabriel-Robez et al. 1986) and fowl (Kaelbling and Fechheimer 1985). (2) Asynapsis of the inverted regions followed by heterosynapsis found in the sand rat (Ashley et al. 1981) and in human (Saadallah and Hulten 1986). (3) Inverted region heterosynapsis occurring in early pachytene, revealed in the deer mouse (Greenbaum and Reed 1984) and in human (Chandley et al. 1987). (4) Homosynapsed inversion loops remaining throughout pachytene found in mice doubly heterozygous for overlapping inversions (Chandley 1982) and in human (Batanian and Hulten 1987). The last type of synapsis is also characteristic of inversions in maize (Maguire 1981; Anderson et al. 1988). Leaving aside the interspecific differencies in synaptic pattern, it is important to find the causes of intraspecific differences for this trait. Do they depend upon the genotype, size or position of the inverted segment? The present paper describes results of pachytene analysis carried out on male mice singly and doubly heterozygous for three overlapping rearrangements in chromosome 1 : two paracentric inversions ln(1)lRk and In(1)12Rk and inversion In(1)lIcg. The latter contains an inverted duplication of the HSR insertion. Parts of the duplication are separated by an inverted euchromatic segment, Electron microscopic (EM) analysis of the synaptonemal complexes (SCs) reveals that synapsis and synaptic adjustment are dependent on the size and location of the inversions and their interaction. The synaptic adjustment of the insertion D-loops occurs at the late zygotene/early pachytene stage. Later, the synaptic adjustment of the inversion loops takes place. A delay in adjustent was found for the double heterozygotes In(1)lIcg/ In(1)lRk and In(1)IIcg/In(1)12Rk. A correspondence between the duration of asynapsis and the probability of association of the pair and heteromorphic bivalents was revealed.
366
In12Rk
Materials and methods Male and female mice homozygous for inversions In(1)lRk and In(1)12Rk were kind gifts of Dr. T.H. Roderick from the Jackson Laboratory. In(l)lRk, hereafter called InlRk, is a paracentric inversion about 45 cM in length on chromosome l (Roderick and Hawes 1974). Study of the banding pattern shows the breakpoints to be within or at the distal side of the most proximal light G-band (1B) and inside the light band just distal to the central bright band (1F) (Davisson and Roderick 1973). In(1)12Rk, hereafter called In12Rk, is a paracentric inversion on chromosome l. According to our data the proximal breakpoint is between the E1 and E2 sub-bands and the distal one is just near the telomere. Chromosome I bearing In(l)lIcg, hereafter called IniIcg, was isolated from a feral population of Mus musculus musculus near Novosibirsk (Agulnik et al. 1987). It possesses a complex rearrangement inclusing the insertion Is(HSR;1C5)1Icg, the inverted euchromatic segment between the 1C5 and IE3 sub-bands and insertion Is(HSR;ID)2Icg. This chromosome presumably arose as a result of inversion in chromosome 1 carrying an additional HSR characteristic of M. m. domesticus (Traut et al. 1984). The HSRs were shown to represent amplified and probably rearranged DNA sequencies (Weith et al. 1987). The proximal breakpoint of the inversion was localized inside the HSR segment and the distal one, between the E3 and E4 sub-bands (Agulnik et al. in press). Thus Is(HSR;1D)2Icg can be considered as an inverted duplication of Is(HSR;1C5) IIcg. Figure l shows a schematic representation of the chromosomes involved in the study. The F1 double and single heterozygotes InlIcg/InlRk and InlIcg/Inl2Rk as well as l n l R k / + and In12Rk/ + were produced by crossing InlRk and In12Rk homozygotes respectively with InlIcg heterozygotes. Three 3 month old males of each genotype were used in the study. Somatic karyotypes of individuals were identified in air-dried bone marrow preparations (Evans et al. 1964) followed by C-band staining (Sumner 1972) which allowed the HSR-bearing chromosome to be visualized. Surface-spread SC preparations were made according to the method of Solari (1980), stained with AgNO3 (Howell and Black 1980) and examined and photographed using a JEM model EM (Jeol, Japan) at 80 kV. Prophase substages were identified according to the criteria proposed by Moses (1980). All measurements were made with a mechanical curvimeter on prints of total magnification x 6,000. The endpoints of synapsis inside and outside inversion loops, as well as the midpoints of the loops, were measured from the centromere, These were then given as a decimal fraction of the mean overall length of the two lateral elements of the given SC.
Results
1nllcg/ + A g e n e r a l d e s c r i p t i o n o f the p a i r i n g p a t t e r n in h e t e r o z y gotes for I n l l c g has b e e n p u b l i s h e d earlier ( B o r o d i n et al. 1990). N e i t h e r t y p i c a l i n v e r s i o n l o o p s , n o r associations b e t w e e n the h e t e r o r n o r p h i c b i v a l e n t a n d the X Y b i v a l e n t were observed. A single D - l o o p (Fig. 2 a ) o f v a r i a b l e size was f o u n d in a small f r a c t i o n o f s p e r m a t o cyte nuclei at late z y g o t e n e / e a r l y p a c h y t e n e (Table 1). T h e size o f the l o o p d e p e n d e d u p o n the stage: it g r a d u a l ly d e c r e a s e d f r o m late z y t o g e n e to the e n d o f early p a chytene. T h e a v e r a g e size was 0.18 t h a t o f the s h o r t ( n o r m a l ) axis. A s s h o w n earlier, m i t o t i c c h r o m o s o m e 1 c a r r y i n g the H S R i n s e r t i o n s is 1.5 times l o n g e r then a n o r m a l c h r o m o s o m e 1 ( A g u l n i k et al. 1987). T h e r e -
,lJJJJIJIIIIJJJ JHJIIIIIHIII .o, Inllcg
I
Fig. i. Locations of the inversion breakpoints on the cytological map of chromosome 1 Table 1. Frequency of the different types of synaptic configurations shown in Figure 2 as a percentage of the total number of pachytene nuclei scored, and fraction of bivalents of each type associated with XY bivalents Karyotype
Number of nuclei
Configuration
Percentage Fraction of bivalents of such of this type bivalents associated with XY bivalent
InlIcg/+ InlRk/+
229 88
InlRk/InlIcg
107
Inl2Rk/+
149
Inl2Rk/InlIcg
147
a b c b c d e f d e f
4.3 _+1.3 58.0-+5.2 4.5_+2.2 49.5+_4.8 3.7.+1.8 2.0___1.1 34.8 -/-_3.9 8.1.+2.2 2.0+ 1.1 43.7-+4.1 11.6.+2.6
0 11.8_+4.5 100 5.7.+3.1 75.0_+21.7 0 37.7 -+6.9 83.3.+10.7 0 30.3-+5.2 70.6.+11.1
fore, the c h r o m a t i n o f the insertions is u n d e r r e p r e s e n t e d in SC structures. T h e m i d p o i n t o f the l o o p is l o c a t e d at a d i s t a n c e o f 0.51 f r o m the c e n t r o m e r e a n d coincides w i t h the p o s i t i o n o f this p o i n t o n the m i t o t i c c h r o m o s o m e m a p (Table 2, Fig. 1). E v e n at early p a c h y t e n e the m a j o r i t y o f nuclei contain o n l y n o r m a l s t r a i g h t bivalents. T h e a v e r a g e size o f the l o n g e s t b i v a l e n t ( p r e s u m a b l y b i v a l e n t 1) m e a s u r e d in h e t e r o z y g o t e s a n d n o r m a l a n i m a l s ( 1 2 . 0 1 + 0 . 4 5 ~tm a n d 1 1 . 7 1 + 0 . 1 5 Bm respectively) does n o t differ ( P > 0,05). H o w e v e r , the l e n g t h o f SC 1 in b o t h is significantly s h o r t e r t h a n in the h o m o z y g o t e s for this a b e r r a t i o n (13.51 + 0 . 4 5 ~tm) ( P < 0 . 0 5 ) . These d a t a s h o w t h a t syna p t i c a d j u s t m e n t o f the h e t e r o m o r p h i c b i v a l e n t in hetero z y g o t e s is o b t a i n e d b y s h o r t e n i n g o f l a t e r a l e l e m e n t segments which a r e u n p a i r e d at e a r l y p a c h y t e n e .
I n l R k / + and I n l R k / I n l l c g
These g e n o t y p e s d i s p l a y i n v e r s i o n l o o p c o n f i g u r a t i o n s in m o r e t h a n h a l f the p a c h y t e n e nuclei (Table 1). M o s t are t y p i c a l i n v e r s i o n l o o p s w i t h i n b i v a l e n t s w h o s e ends are p a i r e d (Fig. 2 b). H o w e v e r , b i v a l e n t s w i t h a s y n a p s e d a n d m o r e e l e c t r o n dense t e l o m e r i c ends to the lateral elements were f o u n d in a few late p a c h y t e n e nuclei
367
Fig. 2 a-L Electron micrographs of silver-stained heteromorphic bivalents of chromosome 1 in male mice heterozygous for different inversions, a InlIcg/+; b, e InlRk/+ and InlRk/InlIcg; d, e, f Inl2Rk/+ and Inl2Rk/InlIcg. A centromere, B proximal end-
point of synaptonemal complex (SC) outside a loop, C proximal endpoint of SC in a loop, D midpoint of SC in a loop, E distal endpoint of SC in a loop, F distals endpoint of SC outside a loop, G telomere, XY sex bivalent
(Fig. 2c). The latter configuration always forms loose or close contact with the sex pair, while the former shows association only in about 12% nuclei (Table 1). Double inversion loops were never found in double heterozygores, despite the fact that the inversions overlapped (Fig. 1). The positions of the endpoints and midpoints of the inversion loop in single heterozygotes are in good agreement with those reported earlier for this karyotype (Moses et al. 1982). Double heteropzygotes differ from single ones by a more distal position of the distal endpoint o f the SC (point F, Fig. 2b) and greater distance
between proximal (B) and distal (F) endpoints of the SC ( P < 0.05, Table 2). The length ratios of the long/short axes in the configuration mentioned above were equal for the two karyotypes (1.12 + 0.03).
Inl2Rk/ ÷ and Inl2Rk/Inllcg Light microscopy did not reveal any heteromorphic bivalents in single heterozygotes for In12Rk at any stages of prophase (Chandley 1982). Our EM study however
368 Table 2. Relative distance from centromere to the endpoints and midpoints of the SC inside and outside the loops in the different synaptic configurations shown in Figure 2 Karyotype
InlRk/+ InlRk/+ InlRk/In] Icg In12Rk/+ Inl2Rk/InlIcg
Configuration a b b e f e f
Number of bivalents 6 34 30 31 7 29 10
Distance to point B
C
D
E
0.41 +0.03 0.20_+0.01 0.18_+0.01 0.52_+0.01 0.74-t-0.03 0.53+0.02 0.76+0.02
0.26_+0.01 0.25-t-0.01 0.64-t-0.01 0.68_+0.01 -
0.51 _+0.02 0.40_+0.02 0.41 +0.02 0.73±0.01
0.54_+0.02 0.59_+0.02 0.81_+0.01a
0.77+0.02 -
-
F
0.60 + 0.03 0.61 -t-0.02" 0.68+0.02 a
0.86+0.01"
" Differences in the point position between single and double heterozygotes are significant (P < 0.05)
allowed us to find three types of heteromorphic configuration c o m m o n for single and double heterozygotes. Partial inversion loops with fully paired ends of axes (Fig. 2d) were found in a small fraction of late zygotene nuclei (Table 1) but the more frequent configuation at all substages of pachytene was the partial inversion loop with unpaired distal segments of the axes (Fig. 2e). A quantitative analysis revealed a significant difference between double and single heterozygotes in respect of the position of the SC endpoints in the loop (points C and E, Fig. 2e). In the former these points were shifted to the telomere (P
CHROMOSOMA © Springer-Verlag 1990
Synapsis in single and double heterozygotes for partially overlapping inversions in chromosome 1 of the house mouse Pavel M. Borodin, Ivan P. Gorlov, and Tatyana Yu. Ladygina Institute of Cytology and Genetics, USSR Academy of Sciences, Siberian Department, Novosibirsk 630090, USSR ReceivedJanuary 16, 1990/in revised form March 4, 1990 Accepted April 11, 1990 by H.C. Macgregor
Abstract. Electron microscopic (EM) analysis of synaptonemal complexes (SC) in single and double heterozygotes for the partially overlapping inversions In0)lIcg, In(l)lRk and In(1)12Rk in chromosome 1 of the house mouse reveals that synapsis and synaptic adjustment are dependent on the size and location of the inversions and interaction between the latter. In(1)lIcg contains insertions of the inverted repeats Is(HSR; 1C 5) lIcg and Is(HSR;1D)2Icg and an inverted euchromatic region. Synaptic adjustment of the D-loops by shortening of the asynapsed segments of the lateral elements belonging to the insertions occurs at the late zytogene to early pachytene stage. Synaptic adjustment of the inversion loops takes place at early to late pachytene. A delay in adjustment was found in the double heterozygotes ln(1)lIcg/In(1)lRk and In(1)lIcg/In(1)12Rk. A correspondence between the lifespan of asynapsis in inverted regions and the probability of association of XY and heteromorphic bivalents was revealed. Introduction Using a surface-spreading technique, loop formation and its resolution at the pachytene stage in male mice heterozygous for the inversions In(1)lRk and In(2)5Rk (Poorman et al. 1981a; Moses et al. 1982) and tandem duplication in chromosome 7 (Poorman et al. 1981b; Moses and Poorman 1981) has been described. Similar observations have been reported in heterozygotes of an inverted duplication of a chromosome 1 homogeneously staining region (HSR) (Borodin et al. 1990). The process of loop resolution connected with partial desynapsis and followed by non-homologous resynapsis has been called "synaptic adjustment" (Moses et al. 1982). It has been suggested that synaptic adjustment is a general phenomenon among different species (Moses 1980). However further studies in inversion heterozygotes have revealed that the fate of the loop during meiotic prophase may vary depending on the species and the type of inversion. Offprint requests to: P.M. Borodin
Anderson et al. (1988) distinguish four types of synaptic behaviour of inversion loops: (1) homosynapsis replaced by heterosynapsis. In addition to the above studies this kind of synaptic adjustment is displayed in mice (Chandley 1982; Tease and Fisher 1986), human (Gabriel-Robez et al. 1986) and fowl (Kaelbling and Fechheimer 1985). (2) Asynapsis of the inverted regions followed by heterosynapsis found in the sand rat (Ashley et al. 1981) and in human (Saadallah and Hulten 1986). (3) Inverted region heterosynapsis occurring in early pachytene, revealed in the deer mouse (Greenbaum and Reed 1984) and in human (Chandley et al. 1987). (4) Homosynapsed inversion loops remaining throughout pachytene found in mice doubly heterozygous for overlapping inversions (Chandley 1982) and in human (Batanian and Hulten 1987). The last type of synapsis is also characteristic of inversions in maize (Maguire 1981; Anderson et al. 1988). Leaving aside the interspecific differencies in synaptic pattern, it is important to find the causes of intraspecific differences for this trait. Do they depend upon the genotype, size or position of the inverted segment? The present paper describes results of pachytene analysis carried out on male mice singly and doubly heterozygous for three overlapping rearrangements in chromosome 1 : two paracentric inversions ln(1)lRk and In(1)12Rk and inversion In(1)lIcg. The latter contains an inverted duplication of the HSR insertion. Parts of the duplication are separated by an inverted euchromatic segment, Electron microscopic (EM) analysis of the synaptonemal complexes (SCs) reveals that synapsis and synaptic adjustment are dependent on the size and location of the inversions and their interaction. The synaptic adjustment of the insertion D-loops occurs at the late zygotene/early pachytene stage. Later, the synaptic adjustment of the inversion loops takes place. A delay in adjustent was found for the double heterozygotes In(1)lIcg/ In(1)lRk and In(1)IIcg/In(1)12Rk. A correspondence between the duration of asynapsis and the probability of association of the pair and heteromorphic bivalents was revealed.
366
In12Rk
Materials and methods Male and female mice homozygous for inversions In(1)lRk and In(1)12Rk were kind gifts of Dr. T.H. Roderick from the Jackson Laboratory. In(l)lRk, hereafter called InlRk, is a paracentric inversion about 45 cM in length on chromosome l (Roderick and Hawes 1974). Study of the banding pattern shows the breakpoints to be within or at the distal side of the most proximal light G-band (1B) and inside the light band just distal to the central bright band (1F) (Davisson and Roderick 1973). In(1)12Rk, hereafter called In12Rk, is a paracentric inversion on chromosome l. According to our data the proximal breakpoint is between the E1 and E2 sub-bands and the distal one is just near the telomere. Chromosome I bearing In(l)lIcg, hereafter called IniIcg, was isolated from a feral population of Mus musculus musculus near Novosibirsk (Agulnik et al. 1987). It possesses a complex rearrangement inclusing the insertion Is(HSR;1C5)1Icg, the inverted euchromatic segment between the 1C5 and IE3 sub-bands and insertion Is(HSR;ID)2Icg. This chromosome presumably arose as a result of inversion in chromosome 1 carrying an additional HSR characteristic of M. m. domesticus (Traut et al. 1984). The HSRs were shown to represent amplified and probably rearranged DNA sequencies (Weith et al. 1987). The proximal breakpoint of the inversion was localized inside the HSR segment and the distal one, between the E3 and E4 sub-bands (Agulnik et al. in press). Thus Is(HSR;1D)2Icg can be considered as an inverted duplication of Is(HSR;1C5) IIcg. Figure l shows a schematic representation of the chromosomes involved in the study. The F1 double and single heterozygotes InlIcg/InlRk and InlIcg/Inl2Rk as well as l n l R k / + and In12Rk/ + were produced by crossing InlRk and In12Rk homozygotes respectively with InlIcg heterozygotes. Three 3 month old males of each genotype were used in the study. Somatic karyotypes of individuals were identified in air-dried bone marrow preparations (Evans et al. 1964) followed by C-band staining (Sumner 1972) which allowed the HSR-bearing chromosome to be visualized. Surface-spread SC preparations were made according to the method of Solari (1980), stained with AgNO3 (Howell and Black 1980) and examined and photographed using a JEM model EM (Jeol, Japan) at 80 kV. Prophase substages were identified according to the criteria proposed by Moses (1980). All measurements were made with a mechanical curvimeter on prints of total magnification x 6,000. The endpoints of synapsis inside and outside inversion loops, as well as the midpoints of the loops, were measured from the centromere, These were then given as a decimal fraction of the mean overall length of the two lateral elements of the given SC.
Results
1nllcg/ + A g e n e r a l d e s c r i p t i o n o f the p a i r i n g p a t t e r n in h e t e r o z y gotes for I n l l c g has b e e n p u b l i s h e d earlier ( B o r o d i n et al. 1990). N e i t h e r t y p i c a l i n v e r s i o n l o o p s , n o r associations b e t w e e n the h e t e r o r n o r p h i c b i v a l e n t a n d the X Y b i v a l e n t were observed. A single D - l o o p (Fig. 2 a ) o f v a r i a b l e size was f o u n d in a small f r a c t i o n o f s p e r m a t o cyte nuclei at late z y g o t e n e / e a r l y p a c h y t e n e (Table 1). T h e size o f the l o o p d e p e n d e d u p o n the stage: it g r a d u a l ly d e c r e a s e d f r o m late z y t o g e n e to the e n d o f early p a chytene. T h e a v e r a g e size was 0.18 t h a t o f the s h o r t ( n o r m a l ) axis. A s s h o w n earlier, m i t o t i c c h r o m o s o m e 1 c a r r y i n g the H S R i n s e r t i o n s is 1.5 times l o n g e r then a n o r m a l c h r o m o s o m e 1 ( A g u l n i k et al. 1987). T h e r e -
,lJJJJIJIIIIJJJ JHJIIIIIHIII .o, Inllcg
I
Fig. i. Locations of the inversion breakpoints on the cytological map of chromosome 1 Table 1. Frequency of the different types of synaptic configurations shown in Figure 2 as a percentage of the total number of pachytene nuclei scored, and fraction of bivalents of each type associated with XY bivalents Karyotype
Number of nuclei
Configuration
Percentage Fraction of bivalents of such of this type bivalents associated with XY bivalent
InlIcg/+ InlRk/+
229 88
InlRk/InlIcg
107
Inl2Rk/+
149
Inl2Rk/InlIcg
147
a b c b c d e f d e f
4.3 _+1.3 58.0-+5.2 4.5_+2.2 49.5+_4.8 3.7.+1.8 2.0___1.1 34.8 -/-_3.9 8.1.+2.2 2.0+ 1.1 43.7-+4.1 11.6.+2.6
0 11.8_+4.5 100 5.7.+3.1 75.0_+21.7 0 37.7 -+6.9 83.3.+10.7 0 30.3-+5.2 70.6.+11.1
fore, the c h r o m a t i n o f the insertions is u n d e r r e p r e s e n t e d in SC structures. T h e m i d p o i n t o f the l o o p is l o c a t e d at a d i s t a n c e o f 0.51 f r o m the c e n t r o m e r e a n d coincides w i t h the p o s i t i o n o f this p o i n t o n the m i t o t i c c h r o m o s o m e m a p (Table 2, Fig. 1). E v e n at early p a c h y t e n e the m a j o r i t y o f nuclei contain o n l y n o r m a l s t r a i g h t bivalents. T h e a v e r a g e size o f the l o n g e s t b i v a l e n t ( p r e s u m a b l y b i v a l e n t 1) m e a s u r e d in h e t e r o z y g o t e s a n d n o r m a l a n i m a l s ( 1 2 . 0 1 + 0 . 4 5 ~tm a n d 1 1 . 7 1 + 0 . 1 5 Bm respectively) does n o t differ ( P > 0,05). H o w e v e r , the l e n g t h o f SC 1 in b o t h is significantly s h o r t e r t h a n in the h o m o z y g o t e s for this a b e r r a t i o n (13.51 + 0 . 4 5 ~tm) ( P < 0 . 0 5 ) . These d a t a s h o w t h a t syna p t i c a d j u s t m e n t o f the h e t e r o m o r p h i c b i v a l e n t in hetero z y g o t e s is o b t a i n e d b y s h o r t e n i n g o f l a t e r a l e l e m e n t segments which a r e u n p a i r e d at e a r l y p a c h y t e n e .
I n l R k / + and I n l R k / I n l l c g
These g e n o t y p e s d i s p l a y i n v e r s i o n l o o p c o n f i g u r a t i o n s in m o r e t h a n h a l f the p a c h y t e n e nuclei (Table 1). M o s t are t y p i c a l i n v e r s i o n l o o p s w i t h i n b i v a l e n t s w h o s e ends are p a i r e d (Fig. 2 b). H o w e v e r , b i v a l e n t s w i t h a s y n a p s e d a n d m o r e e l e c t r o n dense t e l o m e r i c ends to the lateral elements were f o u n d in a few late p a c h y t e n e nuclei
367
Fig. 2 a-L Electron micrographs of silver-stained heteromorphic bivalents of chromosome 1 in male mice heterozygous for different inversions, a InlIcg/+; b, e InlRk/+ and InlRk/InlIcg; d, e, f Inl2Rk/+ and Inl2Rk/InlIcg. A centromere, B proximal end-
point of synaptonemal complex (SC) outside a loop, C proximal endpoint of SC in a loop, D midpoint of SC in a loop, E distal endpoint of SC in a loop, F distals endpoint of SC outside a loop, G telomere, XY sex bivalent
(Fig. 2c). The latter configuration always forms loose or close contact with the sex pair, while the former shows association only in about 12% nuclei (Table 1). Double inversion loops were never found in double heterozygores, despite the fact that the inversions overlapped (Fig. 1). The positions of the endpoints and midpoints of the inversion loop in single heterozygotes are in good agreement with those reported earlier for this karyotype (Moses et al. 1982). Double heteropzygotes differ from single ones by a more distal position of the distal endpoint o f the SC (point F, Fig. 2b) and greater distance
between proximal (B) and distal (F) endpoints of the SC ( P < 0.05, Table 2). The length ratios of the long/short axes in the configuration mentioned above were equal for the two karyotypes (1.12 + 0.03).
Inl2Rk/ ÷ and Inl2Rk/Inllcg Light microscopy did not reveal any heteromorphic bivalents in single heterozygotes for In12Rk at any stages of prophase (Chandley 1982). Our EM study however
368 Table 2. Relative distance from centromere to the endpoints and midpoints of the SC inside and outside the loops in the different synaptic configurations shown in Figure 2 Karyotype
InlRk/+ InlRk/+ InlRk/In] Icg In12Rk/+ Inl2Rk/InlIcg
Configuration a b b e f e f
Number of bivalents 6 34 30 31 7 29 10
Distance to point B
C
D
E
0.41 +0.03 0.20_+0.01 0.18_+0.01 0.52_+0.01 0.74-t-0.03 0.53+0.02 0.76+0.02
0.26_+0.01 0.25-t-0.01 0.64-t-0.01 0.68_+0.01 -
0.51 _+0.02 0.40_+0.02 0.41 +0.02 0.73±0.01
0.54_+0.02 0.59_+0.02 0.81_+0.01a
0.77+0.02 -
-
F
0.60 + 0.03 0.61 -t-0.02" 0.68+0.02 a
0.86+0.01"
" Differences in the point position between single and double heterozygotes are significant (P < 0.05)
allowed us to find three types of heteromorphic configuration c o m m o n for single and double heterozygotes. Partial inversion loops with fully paired ends of axes (Fig. 2d) were found in a small fraction of late zygotene nuclei (Table 1) but the more frequent configuation at all substages of pachytene was the partial inversion loop with unpaired distal segments of the axes (Fig. 2e). A quantitative analysis revealed a significant difference between double and single heterozygotes in respect of the position of the SC endpoints in the loop (points C and E, Fig. 2e). In the former these points were shifted to the telomere (P
