Clinical Generics 1979: 16: 340-346
Interstitial deletion 13q33 resulting from maternal insertional translocation B. S. EMANUEL~,~, E. H. ZACKAI~,~, L. MOREAU',P. COATES~ AND E. ORRECHIO' Departments of 1 Pediatrics and 2 Human Genetics of the University of Pennsylvania School of Medicine, Children's Hospital of Philadelphia, Philadelphia, Pa., U. S . A. A 32-month-old female with a unique interstitial deletion of 13q is presented, including cytogenetic and gene marker studies. The deleted 13 in the patient is a result of malsegregation of a maternal insertional translocation involving chromosomes 7 and 13, 46,XX,ins (7;13)(q22;q32q34). The demonstration of two esterase D alleles in this patient excludes band 13q33 as the! site of the ESD locus, previously assigned to the distal long arm of chromosome 13. The BUdR dye studies reveal that the replicative pattern for 13q31 and 13q21 is not altered by deletion of 13q33 and permit precise delineation of the breakpoints of the rearrangement. Rcceived 5 April, accepted for publication 26 A p r i l 1979
K e y words: Chromosome 13; developmental delay; esterase D; insertional translocation; interstitial deletion.
The literature dcaling with specific terminal deletions of 13q is extensive (Niebuhr 1977). A small number of interstitial deletions of chromosome 13 have also been identified (Noel et al. 1976, Nielsen et al. 1977, Yunis & Ramsay 1978, Wilson et al. 1973, 1977, Orye et al. 1974, Knudson et a1 1976, Francke 1976, Howard et al. 1974). Interstitial deletions are presumably due to 2break events, and have a frequency of 1/500 newborns (Jacobs et al. 1974). It has been estimated that insertions and other 3break rearrangements occur at about 1/10 the frequency of 2-break rearrangements. Several interchromosal insertions have been reported (Chudley et al. 1974, Rethore et al. 1972, Grace et al. 1972, Shapiro & Warburton 1972, Berger et al. 1974, Dennis et al. 1978, Couturier et al. 1977). This is the report of a unique interstitial deletion of 13q, a consequence of malsegregation of a parental insertional transloca0oO9-9163/79/110340-07$02.50/0
tion involving chromosomes 7 and 13. Precise delineation of the breakpoints of this deletion provided the opportunity to examine clinical, gene marker, and chromosomal DNA replicative manifestations associated with deletion of band 13q33. Material and Methods
Peripheral blood leukocytes were grown for 72 h in McCoy's modified 5a medium, supplemented with 15 70 fetal calf serum and PHA. BUdR (Gibco) was added t o the medium of some of the cultures to achieve a final concentration of 200 pg/ml. Colchicine was added 1 h prior to termination of the cultures, and the cells were harvested in the standard manner. The cultures containing BUdR were harvested in subdued light. Airdried chromosome preparations were made. Slides prepared from the leukocytes cul0 1979 Munksgaard, Copenhagen
INTERSTITIAL DELETION 13q33
341
Fig. 1. Patient, age 32 months. Note frontal bossing, hypertelorisrn, upslanting palpebral fissures, and large
ears.
tured without BUdR were G-banded using trypsin pretreatment. Slides from BUdRcontaining cultures were stained with 33258 Hoechst dye, exposed to visible light and subsequently stained with Giemsa as previously described (Emanuel 1978). A minimum of 25 BUdR substituted metaphases per patient were analyzed and scored for late replication or decreased Giemsa stainability of bands 13q21 and 13q31 as compared to the three late replicating bands on the no. 4 chromosomes in the same spreads (p15, q13, q28). This comparison serves as an internal control for BUdR incorporation within a given cell. Starch gel electrophoresis was carried out according to the methods of Coates et al. (Coates et al. 1975), using 0.2 M trismaleate, pH 7.2, bridge buffer and a 1/10 dilution of this buffer in the gel. 13 70 (w/v) gels were electrophoresed at 4 v/cm
overnight at 4°C. Staining for esterase D was according to Hopkinson et al. (Hopkinson et al. 1973), using 4-methylumbelliferyl acetate at pH 6.5. Case Report
The patient, a white female, was evaluated at age 32 months because of hypertelorism and developmental delay (Fig. 1). She was the 3520 g product of a fullterm pregnancy, born to a 26-year-old G,P, mother and a 27-year-old father. Apgar score was 8 following a mid-forceps delivery (used for rotation from posterior position). The child was noted to have hypertelorism, a poor suck, and was lethargic in the neonatal period. Her head and length were at the 90th percentile (father’s head circumference > 90 YO). Alternating esotropia, mild spastic di-
342
EMANUEL ET AL.
plegia and developmental delay were evident at age 6 months. Subsequent developmental evaluation at age 26 months showed marked retardation (9 month motor, 7 month language). Neurologic follow up at that time again demonstrated the mild spastic diplegia. The patient’s growth (both head circumference and length) remained along the 90th percentile. The esotropia was corrected by surgery. Recurrent otitis media has resulted in a bilateral conductive hearing loss. On physical examination at 32 months the patient’s head circumference and length were at the 98th YO. She has true hypertelorism with an interpupillary distance of 6.2 cm (> 97 Yo). Frontal bossing, a prominent sagittal ridge, and upslanting palpebral fissures were noted. The patient has large, normalset ears with overdeveloped lobules. There was a small hemangioma on the left upper chest. The nipples were normally spaced but were hypoplastic. The patient exhibited no speech, cannot walk, but does stand with support. There was a mild hypotonia and weakness in the upper extremities and increased reflexes and weakness in the lower extremities. There was a mild intention tremor in the upper extremities. There was a distally placed triradius on the left palm. The remainder of the dermatoglyphics and physical examination were normal. Results
Cytogenetic Results Trypsin G-banding studies. The proband’s karyotype was initially designated as 46, XX,del(l3)(q32+qter) on the basis of length and G-band pattern of one of her no. 13 chromosomes in all cells examined (Fig. 2). Her phenotypically normal mother has a similarly deleted chromosome 13. Analysis
A
B
dd(13)
U
Fig. 2. Full and partial G-banded karyotypes of patient. A. Arrow indicates chromosome 13 with deletion. B. Chromosome 13 pairs from additional metaphases. The del(13) is placed to the left.
of maternal late prophase chromosomes reveals that the deleted material is inserted into a chromosome 7 (Fig. 3), making the mother’s presumptive karyotype either 46, XX,ins (7;13)(q22;q32q34) or ins (7;13) (q22;q31q32). The G-banding pattern does not permit discrimination between a direct or an inverted insertion, nor does it precisely identify the breakpoints on the no. 13. Chromosome studies of the father and maternal grandparents of the proband were normal. BUdR dye studies. BUdR-substituted metaphase preparations of the propositus, her parents, and maternal grandparents were analyzed for late replication of bands 13q21 and 13q31. The deleted chromosome of the propositus has two bands which replicate late (Fig. 4A). These bands are taken to
INTERSTITIAL DELETION 13q33
7
der(7)
13 dci(13)
343
mother’s karyotype is established as 46,XX, ins(7;13)(q22;q32q34), making the proband’s 46,XX,del( 1 3)(q32-+q34). The frequency of the late replicative pattern for the chromosomes 13 of the propositus and her mother was studied and compared to that for laboratory controls. The replicative pattern for the del( 13) chromosome in the proband and in her mother does not differ from that seen in karyologically normal controls. The percentage of 13q bands in proband and mother which do not late replicate is the same as the laboratory mean established from similar analysis of 20 controls (proband 7 00,mother 3 00, control 5.86 70, S. D. = 2.4 00). The value for their chromosome 4 bands is also within moththe laboratory range (proband 4.7 00, control 11.96 00, s. D. = 4.9 00). er 4.1 00,
Fig. 3. Partial G-banded karyotypes of the patient’s mother showing chromosomes 7 and 13. The arrows on the der(7) indicate the Giemsa positive band derived from the del(13). The arrows on the normal 13s indicate the location of this inserted material.
correspond to bands 13q21 and 13q31. The more distal band, 13q31, is comparable in size to the corresponding band of her normal homologue and of chromosome 13 pairs from karyotypically normal family members. The banding pattern of her del(13) is consistent with loss of band 13q33 with breakpoints in q32 and q34. The banding pattern of the mother’s chromosome 7 and 13 pairs permits confirmation of breakpoints within bands 13q32 and 13q34 (Fig. 4B). It is apparent that the telomeric end of the 13 is present on the deleted 13 and the darker staining band 13q33 is removed and inserted into the no. 7 chromosome (Fig. 4C). The inserted material is not late replicating, eliminating the possible maternal karyotype of 46,XX,ins(7; 13)(q22;q3lq32). Thus the
Fig. 4. BUdR substituted, Hoechst plus Giemsa stained partial karyotypes. A. Chromosome 13 pairs from the patient. B. Chromosome 13 pairs from her mother. C. Maternal chromosomes 7 and 13 with breakpoints indicated on normal homologues (top row). Reconstruction of insertion (bottom row) produced by cutting at breakpoints. Actual der(7) and del(13) present for comparison
344
EMANUEL ET AL.
Enzyme Results There are three commonly occurring electrophoretic phenotypes of human esterase D, Es-DI, Es-D2-1, and Es-D2 which are determined by two alleles, Es-D’ and Es-D’. The father shows phenotype Es-D2-I, the mother shows phenotype Es-D1 and the proband is a heterozygote, Es-D2-1. Discussion
Review of clinical and cytogenetic findings of patients with deletions of 13q has prompted their separation into three groups (Niebuhr 1977). Patients with de1(13)(q33-+ qter) have severe retardation, microcephaly, frontal bossing, “Greek profile” (due to absence of defined nasal bridge), hypertelorism, forward slanting of superior incisors, and large ears with deep sulci helici. The second group, patients with del( 13)(q31,32+ qter) have the additional findings of hypoplastic or absent thumbs, agenesis of 1st metacarpal bone, and fusion of the 4th and 5th metacarpals. Male genital malformations and trigonocephaly are observed more frequently with this latter group. The third group of patients are those with retinoblastoma and relatively few associated congenital defects. Evaluation of several such patients has led to the tentative association of retinoblastoma with interstitial deletion of the distal half of 13q14 (Yunis & Ramsay 1978). Our patient with interstitial deletion 13q33 is severely retarded, with hypertelorism, frontal bossing, and large ears with overdeveloped lobules - features described with de1(13)(q33+qter). She does not have microcephaly and the typical profile of this deletion. The absence of abnormal extremities and retinoblastoma in our patient is consistent with previous reports associating these findings with deletions of other segments of 13q. The locus for esterase D (ESD) has been
previously assigned to the distal half of the long arm of chromosome 13 (Robson et al. 1976). The demonstration of two ESD alleles in the proband (ESDI and ESD2) with a demonstrable deletion of band 13q33 excludes this as the site of the ESD locus, and further helps in the precise mapping of this locus. The proband received a del(13) as a consequence of malsegregation of one of the derivative chromosomes involved in a maternal insertional translocation, 46,XX,ins (7;13)(q22;q32q34). The most likely meiotic configuration for such a rearrangement would consist of two unequal bivalents. One would expect that these bivalents would segregate at random, resulting in equal numbers of gametes carrying either the normal 7 and 13, the der(7) and der(l3), or either of the two derivative chromosomes with the complementary normal chromosome. Thus, this translocation heterozygote mother is at risk for production of abnormal, viable offspring partially trisomic or monosomic for the segment 13q32+q34. There are seven families in the literature (Chudley et al. 1974, RethorC et al. 1972, Grace et al. 1972, Shapiro & Warburton 1972, Berger et al. 1974, Dennis et al. 1978, Couturier et al. 1977) with an interchromosomal insertion giving rise to unbalanced offspring. In only two families (Chudley et al. 1974, Dennis et al. 1978) were males the carriers of the insertional rearrangement. Among all the balanced carriers, including those of the present report, there are 10 females and 4 males. The significance of this sex difference may become more apparent when additional rearrangements of this type are detected and studied. The BUdR dye studies have revealed that the pattern of late replication for 13q31 and 13q21 is not altered by the deletion of 13q33 and the material contiguous to it in q32 and q34. Since the integrity of the replicative pattern is not disturbed by the
INTERSTITIAL DELETION 13q33
insertional rearrangement between chromosomes 7 a n d 13, it appears that the controls for replicative timing of chromosome 13 reside within the individual bands. We have previously reported (Emanuel 1978) that other alterations in t h e position a n d organization of chromosomal regions do n o t affect their replicative pattern. A combination of the standard banding techniques was inadequate for precise delineation of the breakpoints cf this rearrangement. Use of B U d R incorporation late in S phase and the B U d R dye technique enabled us to distinguish between a terminal and a n interstitial deletion as well as t o define the breakpoints precisely. This case study emphasizes how t h e combination of refined cytogenetic techniques and qualitative gene marker studies i n unique deletions can add m u c h to the growing pool of information about the gene localization, and phenotypic expression of specific chromosomal segments.
Acknowledgments
T h e authors wish t o acknowledge the assistance of Mi-. J. Loftus, Ms. S. Melicia, and Ms. D. Eunpu. References
Berger, R., G. Touati, J. Derre, M. A. Ortiz & J. Martinetti (1974). &ri d u Chats syndrome with maternal insertional translocation. Clin. Genet. 5, 428-432. Chudley, A. E., F. Bauder, M. Ray, P. J. McAlpine, S.D. J. Pena & J.L. Hamerton (1974). Familial mental retardation in a family with an inherited chromosome rearrangement. J . med. Genet. 11, 353-363. Coates, P. M., M. A. Mestriner & D. A. Hopkinson (1975). A preliminary genetic interpretation of the esterase isozymes of human tissues. Ann. hum. Genet. 39, 1-20. Couturier, J., A. Aurias, M. Prieur & A. Barois (1977). Trisomie partielle pour le bras long d u chromosome 2 par rnalsCgrCgation d’une insertion maternelle: ins (6;2)(p22;q24q34). A n n . Ge‘nPt. 20, 52-55.
345
Dennis, N.R., R.L. Neu & R.M. Bannerman (1978). Duplication 2q33+2q37 due to paternal ins (12;2) translocation. Amer. J . med. Genet. 1, 271-277. Emanuel, B. S. (1978). Compound lateral asymmetry in human chromosome 6: BrdU-Dye studies of 6q12+6q14. Amer. J . hum. Genet. 30, 153-159. Francke, U. (1976). Retinoblastoma and chromosome 13. Birth Defects: Original Article Series, Vol. X I I , No. 7, 131-134. Grace, E., G. R. Sutherland & A. D. Bain (1972). Familial insertional translocation. Lancet ii, 231. Hopkinson, D. A., M. A. Mestriner, J. A. Cortner & H.Harris (1973). Esterase D: a new human polymorphism. Ann. hum. Genet. 37, 119-137. Howard, R.O., W.R. Breg, D . M . Albert & R. L. Lesser (1974). Retinoblastoma and chromosome abnormality - partial deletion of the long arm of chromosome 13. Arch. Ophthalmol. 92, 490-493. Jacobs, P. A., M. Melville, S. Ratcliffe, A. J . Keay & J. Syme (1974). A cytogenetic survey of 11,680 newborn infants. Ann. hum. Genet. 37, 359-376. Knudson, A.G., Jr., A . T . Meadows, W.W. Nichols & R. Hill (1976). Chromosomal deletion and retinoblastoma. New Engl. J . Med. 295, 1120-1123. Niebuhr, E. (1977). Partial trisomies and deletions of chromosome 13. New Chromosomal Syndromes, ed. J . J. Yunis. New York, Academic Press, pp. 273-299. Nielsen, J., A. Homma, F. Christiansen, K. Rasmussen & P. Saldafia-Garcia (1977). Deletion long arm 13. Hum. Genet. 37, 339345. Noel, B., B. Quack & M. Rethork (I976). Partial deletions and trisomies of chromosome 13: mapping of bands associated with particular malformations. Clin. Genet. 9, 593-602. Orye, E., M. J. Delbeke & Vandenabeele (1974). Retinoblastoma and long arm deletion of chromosome 13. Attempts to define the deleted segment. Clin. Genet. 5, 457464. RethorC, M., J. Lejeune, S. Carpentier, M. Prieur, B. Dutrillaux, Ph. Seringe, A. Rossier & J. C. Job (1972). Trisomie pour la partie distale du bras court du chromosome 3 chez trois Germains. Premier exemple d’insertion chromosomique. Ins (7;3) (q31;p21p26). Ann. G i n i t . 15, 159-165. Robson, E. B., D. A. Hopkinson, K. E. Buckton, J. Robinson & P. E. Polani (1976). Family
346
E M A N U E L E T AL
studies on esterase D and chromosome 13 in man. Birth Defects: Original Article Series, Vol. XII, No. 7, 351-354. Shapiro, L. R. & D. Warburton (1972). Interstitial translocation in man. Lancet ii, 713. Wilson, M. G., A. J. Ebbin, J. W. Towner, W. H. Spencer (1977). Chromosomal anomalies in patients with retinoblastoma. Clin. Genet. 12, 1-8. Wilson, M. G., J. W. Towner & A. Fujimoto (1973). Retinoblastoma and D-chromosome deletions. Amer. J . hum. Genet. 25, 57-61.
Yunis, J. J. & N. Ramsay (1978). Retinoblastoma and subband deletion of chromosome 13. Amer. J . Dis. Child 132. 161-163. Address: Beverly S. Emanuel, Ph. D. Cytogenetic Laboratory The Children’s Hospital of Philadelphia 34th and Civic Center Boulevard Philadelphia, Pa. 19104 U.S. A .
Interstitial deletion 13q33 resulting from maternal insertional translocation B. S. EMANUEL~,~, E. H. ZACKAI~,~, L. MOREAU',P. COATES~ AND E. ORRECHIO' Departments of 1 Pediatrics and 2 Human Genetics of the University of Pennsylvania School of Medicine, Children's Hospital of Philadelphia, Philadelphia, Pa., U. S . A. A 32-month-old female with a unique interstitial deletion of 13q is presented, including cytogenetic and gene marker studies. The deleted 13 in the patient is a result of malsegregation of a maternal insertional translocation involving chromosomes 7 and 13, 46,XX,ins (7;13)(q22;q32q34). The demonstration of two esterase D alleles in this patient excludes band 13q33 as the! site of the ESD locus, previously assigned to the distal long arm of chromosome 13. The BUdR dye studies reveal that the replicative pattern for 13q31 and 13q21 is not altered by deletion of 13q33 and permit precise delineation of the breakpoints of the rearrangement. Rcceived 5 April, accepted for publication 26 A p r i l 1979
K e y words: Chromosome 13; developmental delay; esterase D; insertional translocation; interstitial deletion.
The literature dcaling with specific terminal deletions of 13q is extensive (Niebuhr 1977). A small number of interstitial deletions of chromosome 13 have also been identified (Noel et al. 1976, Nielsen et al. 1977, Yunis & Ramsay 1978, Wilson et al. 1973, 1977, Orye et al. 1974, Knudson et a1 1976, Francke 1976, Howard et al. 1974). Interstitial deletions are presumably due to 2break events, and have a frequency of 1/500 newborns (Jacobs et al. 1974). It has been estimated that insertions and other 3break rearrangements occur at about 1/10 the frequency of 2-break rearrangements. Several interchromosal insertions have been reported (Chudley et al. 1974, Rethore et al. 1972, Grace et al. 1972, Shapiro & Warburton 1972, Berger et al. 1974, Dennis et al. 1978, Couturier et al. 1977). This is the report of a unique interstitial deletion of 13q, a consequence of malsegregation of a parental insertional transloca0oO9-9163/79/110340-07$02.50/0
tion involving chromosomes 7 and 13. Precise delineation of the breakpoints of this deletion provided the opportunity to examine clinical, gene marker, and chromosomal DNA replicative manifestations associated with deletion of band 13q33. Material and Methods
Peripheral blood leukocytes were grown for 72 h in McCoy's modified 5a medium, supplemented with 15 70 fetal calf serum and PHA. BUdR (Gibco) was added t o the medium of some of the cultures to achieve a final concentration of 200 pg/ml. Colchicine was added 1 h prior to termination of the cultures, and the cells were harvested in the standard manner. The cultures containing BUdR were harvested in subdued light. Airdried chromosome preparations were made. Slides prepared from the leukocytes cul0 1979 Munksgaard, Copenhagen
INTERSTITIAL DELETION 13q33
341
Fig. 1. Patient, age 32 months. Note frontal bossing, hypertelorisrn, upslanting palpebral fissures, and large
ears.
tured without BUdR were G-banded using trypsin pretreatment. Slides from BUdRcontaining cultures were stained with 33258 Hoechst dye, exposed to visible light and subsequently stained with Giemsa as previously described (Emanuel 1978). A minimum of 25 BUdR substituted metaphases per patient were analyzed and scored for late replication or decreased Giemsa stainability of bands 13q21 and 13q31 as compared to the three late replicating bands on the no. 4 chromosomes in the same spreads (p15, q13, q28). This comparison serves as an internal control for BUdR incorporation within a given cell. Starch gel electrophoresis was carried out according to the methods of Coates et al. (Coates et al. 1975), using 0.2 M trismaleate, pH 7.2, bridge buffer and a 1/10 dilution of this buffer in the gel. 13 70 (w/v) gels were electrophoresed at 4 v/cm
overnight at 4°C. Staining for esterase D was according to Hopkinson et al. (Hopkinson et al. 1973), using 4-methylumbelliferyl acetate at pH 6.5. Case Report
The patient, a white female, was evaluated at age 32 months because of hypertelorism and developmental delay (Fig. 1). She was the 3520 g product of a fullterm pregnancy, born to a 26-year-old G,P, mother and a 27-year-old father. Apgar score was 8 following a mid-forceps delivery (used for rotation from posterior position). The child was noted to have hypertelorism, a poor suck, and was lethargic in the neonatal period. Her head and length were at the 90th percentile (father’s head circumference > 90 YO). Alternating esotropia, mild spastic di-
342
EMANUEL ET AL.
plegia and developmental delay were evident at age 6 months. Subsequent developmental evaluation at age 26 months showed marked retardation (9 month motor, 7 month language). Neurologic follow up at that time again demonstrated the mild spastic diplegia. The patient’s growth (both head circumference and length) remained along the 90th percentile. The esotropia was corrected by surgery. Recurrent otitis media has resulted in a bilateral conductive hearing loss. On physical examination at 32 months the patient’s head circumference and length were at the 98th YO. She has true hypertelorism with an interpupillary distance of 6.2 cm (> 97 Yo). Frontal bossing, a prominent sagittal ridge, and upslanting palpebral fissures were noted. The patient has large, normalset ears with overdeveloped lobules. There was a small hemangioma on the left upper chest. The nipples were normally spaced but were hypoplastic. The patient exhibited no speech, cannot walk, but does stand with support. There was a mild hypotonia and weakness in the upper extremities and increased reflexes and weakness in the lower extremities. There was a mild intention tremor in the upper extremities. There was a distally placed triradius on the left palm. The remainder of the dermatoglyphics and physical examination were normal. Results
Cytogenetic Results Trypsin G-banding studies. The proband’s karyotype was initially designated as 46, XX,del(l3)(q32+qter) on the basis of length and G-band pattern of one of her no. 13 chromosomes in all cells examined (Fig. 2). Her phenotypically normal mother has a similarly deleted chromosome 13. Analysis
A
B
dd(13)
U
Fig. 2. Full and partial G-banded karyotypes of patient. A. Arrow indicates chromosome 13 with deletion. B. Chromosome 13 pairs from additional metaphases. The del(13) is placed to the left.
of maternal late prophase chromosomes reveals that the deleted material is inserted into a chromosome 7 (Fig. 3), making the mother’s presumptive karyotype either 46, XX,ins (7;13)(q22;q32q34) or ins (7;13) (q22;q31q32). The G-banding pattern does not permit discrimination between a direct or an inverted insertion, nor does it precisely identify the breakpoints on the no. 13. Chromosome studies of the father and maternal grandparents of the proband were normal. BUdR dye studies. BUdR-substituted metaphase preparations of the propositus, her parents, and maternal grandparents were analyzed for late replication of bands 13q21 and 13q31. The deleted chromosome of the propositus has two bands which replicate late (Fig. 4A). These bands are taken to
INTERSTITIAL DELETION 13q33
7
der(7)
13 dci(13)
343
mother’s karyotype is established as 46,XX, ins(7;13)(q22;q32q34), making the proband’s 46,XX,del( 1 3)(q32-+q34). The frequency of the late replicative pattern for the chromosomes 13 of the propositus and her mother was studied and compared to that for laboratory controls. The replicative pattern for the del( 13) chromosome in the proband and in her mother does not differ from that seen in karyologically normal controls. The percentage of 13q bands in proband and mother which do not late replicate is the same as the laboratory mean established from similar analysis of 20 controls (proband 7 00,mother 3 00, control 5.86 70, S. D. = 2.4 00). The value for their chromosome 4 bands is also within moththe laboratory range (proband 4.7 00, control 11.96 00, s. D. = 4.9 00). er 4.1 00,
Fig. 3. Partial G-banded karyotypes of the patient’s mother showing chromosomes 7 and 13. The arrows on the der(7) indicate the Giemsa positive band derived from the del(13). The arrows on the normal 13s indicate the location of this inserted material.
correspond to bands 13q21 and 13q31. The more distal band, 13q31, is comparable in size to the corresponding band of her normal homologue and of chromosome 13 pairs from karyotypically normal family members. The banding pattern of her del(13) is consistent with loss of band 13q33 with breakpoints in q32 and q34. The banding pattern of the mother’s chromosome 7 and 13 pairs permits confirmation of breakpoints within bands 13q32 and 13q34 (Fig. 4B). It is apparent that the telomeric end of the 13 is present on the deleted 13 and the darker staining band 13q33 is removed and inserted into the no. 7 chromosome (Fig. 4C). The inserted material is not late replicating, eliminating the possible maternal karyotype of 46,XX,ins(7; 13)(q22;q3lq32). Thus the
Fig. 4. BUdR substituted, Hoechst plus Giemsa stained partial karyotypes. A. Chromosome 13 pairs from the patient. B. Chromosome 13 pairs from her mother. C. Maternal chromosomes 7 and 13 with breakpoints indicated on normal homologues (top row). Reconstruction of insertion (bottom row) produced by cutting at breakpoints. Actual der(7) and del(13) present for comparison
344
EMANUEL ET AL.
Enzyme Results There are three commonly occurring electrophoretic phenotypes of human esterase D, Es-DI, Es-D2-1, and Es-D2 which are determined by two alleles, Es-D’ and Es-D’. The father shows phenotype Es-D2-I, the mother shows phenotype Es-D1 and the proband is a heterozygote, Es-D2-1. Discussion
Review of clinical and cytogenetic findings of patients with deletions of 13q has prompted their separation into three groups (Niebuhr 1977). Patients with de1(13)(q33-+ qter) have severe retardation, microcephaly, frontal bossing, “Greek profile” (due to absence of defined nasal bridge), hypertelorism, forward slanting of superior incisors, and large ears with deep sulci helici. The second group, patients with del( 13)(q31,32+ qter) have the additional findings of hypoplastic or absent thumbs, agenesis of 1st metacarpal bone, and fusion of the 4th and 5th metacarpals. Male genital malformations and trigonocephaly are observed more frequently with this latter group. The third group of patients are those with retinoblastoma and relatively few associated congenital defects. Evaluation of several such patients has led to the tentative association of retinoblastoma with interstitial deletion of the distal half of 13q14 (Yunis & Ramsay 1978). Our patient with interstitial deletion 13q33 is severely retarded, with hypertelorism, frontal bossing, and large ears with overdeveloped lobules - features described with de1(13)(q33+qter). She does not have microcephaly and the typical profile of this deletion. The absence of abnormal extremities and retinoblastoma in our patient is consistent with previous reports associating these findings with deletions of other segments of 13q. The locus for esterase D (ESD) has been
previously assigned to the distal half of the long arm of chromosome 13 (Robson et al. 1976). The demonstration of two ESD alleles in the proband (ESDI and ESD2) with a demonstrable deletion of band 13q33 excludes this as the site of the ESD locus, and further helps in the precise mapping of this locus. The proband received a del(13) as a consequence of malsegregation of one of the derivative chromosomes involved in a maternal insertional translocation, 46,XX,ins (7;13)(q22;q32q34). The most likely meiotic configuration for such a rearrangement would consist of two unequal bivalents. One would expect that these bivalents would segregate at random, resulting in equal numbers of gametes carrying either the normal 7 and 13, the der(7) and der(l3), or either of the two derivative chromosomes with the complementary normal chromosome. Thus, this translocation heterozygote mother is at risk for production of abnormal, viable offspring partially trisomic or monosomic for the segment 13q32+q34. There are seven families in the literature (Chudley et al. 1974, RethorC et al. 1972, Grace et al. 1972, Shapiro & Warburton 1972, Berger et al. 1974, Dennis et al. 1978, Couturier et al. 1977) with an interchromosomal insertion giving rise to unbalanced offspring. In only two families (Chudley et al. 1974, Dennis et al. 1978) were males the carriers of the insertional rearrangement. Among all the balanced carriers, including those of the present report, there are 10 females and 4 males. The significance of this sex difference may become more apparent when additional rearrangements of this type are detected and studied. The BUdR dye studies have revealed that the pattern of late replication for 13q31 and 13q21 is not altered by the deletion of 13q33 and the material contiguous to it in q32 and q34. Since the integrity of the replicative pattern is not disturbed by the
INTERSTITIAL DELETION 13q33
insertional rearrangement between chromosomes 7 a n d 13, it appears that the controls for replicative timing of chromosome 13 reside within the individual bands. We have previously reported (Emanuel 1978) that other alterations in t h e position a n d organization of chromosomal regions do n o t affect their replicative pattern. A combination of the standard banding techniques was inadequate for precise delineation of the breakpoints cf this rearrangement. Use of B U d R incorporation late in S phase and the B U d R dye technique enabled us to distinguish between a terminal and a n interstitial deletion as well as t o define the breakpoints precisely. This case study emphasizes how t h e combination of refined cytogenetic techniques and qualitative gene marker studies i n unique deletions can add m u c h to the growing pool of information about the gene localization, and phenotypic expression of specific chromosomal segments.
Acknowledgments
T h e authors wish t o acknowledge the assistance of Mi-. J. Loftus, Ms. S. Melicia, and Ms. D. Eunpu. References
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