Contiguous Gene Syndromes Due to Deletions in the Distal Short

Proc. Natl. Acad. Sci. USA Vol. 86, pp. 10001-10005, December 1989 Genetics Contiguous gene syndromes due to deletions in the distal short arm of the human X chromosome (steroid sulfatase deficiency/Kalimann syndrome/chondrodysplasia punctata/chromosome X-linked mental retardation/short stature) A. BALLABIO*t, B. BARDONIt, R. CARROZZO§, G. ANDRIA*, D. BICK¶, L. CAMPBELL¶, B. HAMELII M. A. FERGUSON-SMITH**, G. GIMELLItt, M. FRACCAROt, P. MARASCHIOt, 0. ZUFFARDIt, S. GUIOLIt, AND G. CAMERINOT,tt *Department of Pediatrics, University of Reggio Calabria, Catanzaro, Italy; tBiologia e Genetica Medica, University of Pavia, Pavia, Italy; §Department of Pediatrics, University of Naples, Naples, Italy; $University of Texas, Health Center, San Antonio, TX; I1Department of Human Genetics, University of Nijmegen, Nijmegen, The Netherlands; **Department of Pathology, University of Cambridge, Cambridge, United Kingdom; and ttG. Gaslini Institute, Genoa, Italy Communicated by Stanley M. Gartler, September 11, 1989 (received for review August 1, 1989) ABSTRACT Mendelian inherited disorders due to dele- (MRX). The codeletion ofcontiguous genes was postulated to tions of adjacent genes on a chromosome have been described explain the association of these disorders. as "contiguous gene syndromes." Short stature, chondrodys- To confirm this hypothesis and to construct a disease map plasia punctata, mental retardation, steroid sulfatase defi- of the region, we have performed DNA analysis in patients ciency, and Kallmann syndrome have been found as isolated with interstitial and terminal deletions in Xp22-pter and with entities or associated in various combinations in 27 patients X/Y translocations, using cDNA and genomic probes span- with interstitial and terminal deletions involving the distal short ning the region. arm of the X chromosome. The use of cDNA and genomic probes from the Xp22-pter region allowed us to identify 12 different deletion intervals and to confirm, and further refine, MATERIALS AND METHODS the chromosomal assignment of X-linked recessive chondro- Patients. Patients' clinical features and cytogenetic data are dysplasia punctata and Kallmann syndrome genes. A putative summarized in Table 1. Patients have been numbered from 1 pseudoautosomal gene affecting height and an X-linked non- to 16 according to a telomere-centromere order of their specific mental retardation gene have been tentatively assigned proximal breakpoint. to specific intervals. The deletion panel described is a useful Cytogenetic analysis revealed an X/Y translocation, as- tool for mapping new sequences and orienting chromosome sociated with loss of material in the distal Xp, in five patients walks in the region. (patients 1, 2, 8, 9, and 11). Three patients (patients 6, 15, and 16) showed a terminal deletion of the distal Xp. The distal part of the short arm of the X chromosome -is a Patients with X/Y translocations and terminal deletions region of the human genome which shows several very (patients 1, 2, 6, 8, 9, 11, 15, and 16) showed SS, as did all peculiar features. It escapes X inactivation (1), and it shares heterozygous female relatives. The only exception was pa- homology with both the short arm (2-5) and the long arm tient 8, whose height was between the 10th and the 25th (6-9) of the Y chromosome. centile at age 6 yr and 4 mo. However, his first cousin, also In a recent study we have demonstrated a high frequency affected by the same chromosomal abnormality, was below of deletions in this region (10). This high frequency may be the 3rd centile for height when observed at age 6 yr and 6 mo. due to the nonlethal phenotypic effects of these deletions in SS has been described in the original reports of cases 1 (20), the male hemizygous status. A possible role of aberrant 11 (12), 15 (19), and 16 (14). In patients 2, 6, and 9 the height recombination between homologous regions of sex chromo- was 131, 65, and 108 cm at age 13 yr (<3rd centile), 8 mo somes has been proposed (8, 9, 11). (<3rd centile), and 6 yr (5th centile), respectively. The study of interstitial and terminal deletions has enabled Cases 2, 6, 8, 9, 11, 15, and 16 have been diagnosed as the assignment of several genes to Xp22.3-pter. These in- CDPX. All of these patients showed unusual facies charac- clude the following, with their Mendelian Inheritance in Man terized by nasal hypoplasia. Distal phalangeal hypoplasia was (MIM) numbers in parentheses: (i) steroid sulfatase (STS; observed in patients 6, 8, 9, 15, and 16. Epiphyseal stippling MIM 30810) (12), a microsomal enzyme whose deficiency is was documented by x-ray analysis in patients 6, 9, 15, and 16. responsible for X-linked ichthyosis (XLI); (ii) the Xg blood X-ray analysis did not reveal epiphyseal stippling in patients group gene (13); (iii) MIC2 (MIM 31347) (2), a gene coding for 2, 8, and 9 when they were examined at ages 13, 6, and 13, a surface antigen recognized by monoclonal antibody 12E7; respectively. The first cousin of patient 8, affected by the (iv) chondrodysplasia punctata (CDPX; MIM 30295) (14, 15), same chromosomal abnormality, showed tracheal and tarsal a congenital bone dysplasia characterized by unusual face stippling when examined at age 1 mo. Short limbs were due to nasal hypoplasia and by radiologic stippling of epi- observed in patients 2, 9, 11, and 15. physes; and (v) Kallmann syndrome (KAL; MIM 30870) (16), Patients 4, 8, 9, 11, 14, 15, and 16 showed MRX. Patients which is characterized by hypogonadotropic hypogonadism 8 (15), 14 (17), and 16 (14) have already been reported to have and anosmia. variable degrees ofMRX. Patient 4 is the first oftwo brothers In the last few years we and other investigators have with XLI and MRX who were admitted to the clinic because described patients showing complex phenotypes characterized by the association of XLI, due to STS deficiency, with Abbreviations: STS, steroid sulfatase; XLI, X-linked ichthyosis; features of CDPX and KAL (14-19). Some of these patients CDPX, X-linked chondrodysplasia punctata; KAL, Kallmann syn- also showed short stature (SS) and mental retardation drome; SS, short stature; MRX, X-linked nonspecific mental retardation. tPresent address: Institute for Molecular Genetics, Baylor College of The publication costs of this article were defrayed in part by page charge Medicine, One Baylor Plaza, Houston, TX 77030. payment. This article must therefore be hereby marked "advertisement" #4To whom reprint requests should be addressed at: Biologia Gen- in accordance with 18 U.S.C. §1734 solely to indicate this fact. erale e Genetica Medica, Via Forlanini 14, Pavia, Italy. 10001 Downloaded by guest on September 27, 2021 10002 Genetics: Ballabio et al. Proc. Natl. Acad. Sci. USA 86 (1989)

Table 1. Cytogenetic and clinical features of male patients with probes labeled by oligolabeling or nick-translation. Methods deletions in Xp22-pter for DNA extraction and digestion, Southern blotting, probe No. Initials Ref. Karyotype Clinical features labeling, hybridization, and washing were as previously 1 G.C. 20 46,X,t(X;Y); SS described (10, 21). (Xqter- Xp223: Molecular probes used are described in Table 2. STB14 is Ypll-*Yqter) a genomic clone from the STS gene derived from a previously 2 A.M. * 46,Y,der(X),t(X;Y) SS, CDPX isolated STS cDNA clone (28). All probes except G1.3 detect (p22.3;qll) single-copy sequences on the X chromosome at high- 3 A.B. 10 Normal XLI stringency conditions. G1.3 belongs to a family of sequences 4 M.P. 10 Normal XLI, MRX dispersed on the X chromosome and recognizes five different 5 M.C. 10 Normal XLI homologous regions (DFN22SJ-5) (25). Probes MiA, 6 F.A. * 46,Y,del(X)(p22.3) SS, CDPX, (MRX),t XLI STB14, 71-7A, and GMGXY19 recognize homologous se- 7 R.D. 10 Normal XLI quences on Xp22.3 and Yq. 8 M.S. 15 46,Y,der(X)t(X;Y) SS,t CDPX, MRX, XLI (p22.3;qll) 9 F.L. 11 46,Y,der(X)t(X;Y) SS, CDPX, MRX, XLI RESULTS (p22.33;qll.2) DNA analysis results are shown in Table 3. Data from Tables 10 f 10 Normal XLI 1 and 3 have been schematically combined in Fig. 1. 11 A.M. 12 46,Y,der(X)t(X;Y) SS, CDPX, MRX, XLI, Fig. 2 shows the hybridization of probe G1.3 to the DNA (p22.3;qll.2) KAL of seven unrelated patients with deletions in Xp22.3-pter. 12 D.S. 16 Normal XLI, KAL Molecular characterization defines 12 deletion intervals (a to 13 J.G. * Normal XLI, KAL§ 1 in Fig. 1): 14 R.C. 17 Normal XLI, KAL, MRX Interval a spans from the telomere to the breakpoint of 15 J.D. 19 46,Y,del(X)(p22.3) SS, CDPX, MRX, XLI, patient 1. It contains part of the pseudoautosomal region and KAL a putative pseudoautosomal locus affecting height (see Dis- 16 D.N. 14 46,Y,del(X)(p22.3) SS, CDPX, MRX, XLI, cussion). KAL Interval b spans from the breakpoint of patient 1 to the *Cases 2, 6, and 13 are unpublished. breakpoint ofpatient 2. It includes both MIC2 and CDPX loci. tSee patients' description. tIncludes 12 unrelated patients with XLI. Interval c spans from the breakpoint of patient 2 to the §Patient 13 also showed pes cavus and epilepsy. distal breakpoints of patients 4 and 14 and contains the DXS31 locus. of psychomotor delay with I.Q.s of 76 and 64 (assessed by Interval d spans from the distal breakpoints ofpatients 4 and Wechsler test) at ages 9 and 5 yr, respectively. Patient 6 was 14 to the distal breakpoints of patients 5, 7, 10, 12, and 13. A examined at the age of 4 mo, so it will be necessary to assess locus for X-linked nonspecific mental retardation (MRX) his intellectual development at a later date. Patients 9 and 11 could be tentatively assigned to this interval (see Discussion). showed I.Q.s of 66 and 70 at age 5 and 13 yr, respectively. A Intervals e, f, and g are defined by the distal breakpoints of review of the record of case 15 showed that the patient was patients 3, 5, 7, 10, 12, and 13, and by the proximal breakpoints functioning at less than a 3-mo age when tested at 6 mo by the of patients 3, 4, 5, 6, and 7. They contain STS, DYS74, and Denver developmental screening test. DXS237 loci. The order of these three intervals, with respect Patients 3-16 show XLI due to STS deficiency. Twelve to each other, cannot at present be unequivocally established. unrelated patients with isolated XLI showing the same results However, of the six possible orders, two, telgfen and after molecular analysis have been combined under "case 10." tel-cen, can be excluded, since they would imply the Patients 3, 5, 7, and 10 (10) showed XLI as the only clinical presence of two distinct deletions in patients 3, 4, and 5. feature, since SS, MRX, and KAL were carefully ruled out. Interval h spans from the proximal breakpoints of patients Patients 11, 12, 13, 14, 15, and 16 were shown to have 6 and 7 to the proximal breakpoints of patients 8, 9, and 10 KAL. In cases 11, 12, 13, 14, and 16 the diagnosis was made and contains the DFN22SJ locus. on the basis of clinical examination of external genitalia, Interval i spans from the proximal breakpoints of patients endocrinological tests, and evaluation of olfactory function 8, 9, and 10 to the proximal breakpoints ofpatients 11, 12, and (16). Patients 12 (16) and 14 (17) have KAL. Patient 11 has a 13 and contains the KAL locus. deletion of the distal Xp involving the STS gene (12). We Interval j spans from the proximal breakpoints of patients reanalyzed this patient 8 yr later and found that, in addition 11, 12, and 13 to the proximal breakpoint of patient 14 and to CDPX and XLI, he was also affected with hypogonado- contains the DNF22S2 locus. tropic hypogonadism and anosmia. Patient 13 also showed unilateral renal aplasia, pes cavus, and epilepsy. Patient 15 Table 2. Molecular probes used had low gonadotropin levels at 2 mo of age. The patient died X-chromosome of a respiratory infection at 8 mo. Postmortem examination Probe Ref. Locus region revealed complete absence of olfactory bulbs and tracts and unilateral renal hypoplasia (19). Reevaluation of olfactory p2B 22 MIC2 pter function in patient 16, now 13 yr old, was difficult because of M1A 6 DXS31 pter-p22.3 MRX; nevertheless, he appeared to have hyposmia. His STB14 10 STS p22.32 8-yr-old brother, who has the same chromosomal abnormal- GMGXY19 23 DYS74 p22.3 ity, also has hyposmia, a very small phallus, and prepubertal GMGX9 24 DXS237 p22.32 gonadotropin levels. G1.3a 25 DNF22S1 p22.3-p21.2 Direct DNA Analysis. DNA was obtained from patients' (G1.3b) 25 DNF22S2 p22.3-p21.2 venous blood samples, cultured fibroblasts, or human- (G1.3d) 25 DNF22S4 p22.3-p21.2 hamster hybrid cell lines. It was digested with EcoRI or Taq (G1.3e) 25 DNF22S5 p22.3-p21.2 I, Southern blotted on diazobenzyloxymethyl (DBM) or dic56 26 DXS143 p22.3-p22.2 Hybond N (Amersham) paper, and hybridized with DNA 71-7A 27 DXS69 p22.3-p21 Downloaded by guest on September 27, 2021 Genetics: Ballabio et al. Proc. Natl. Acad. Sci. USA 86 (1989) 10003

Table 3. Molecular analysis of deletions in Xp22-pter Patient DXS- D YS- DXS- DNF- DNF- DXS- DNF- DNF- DXS- no. MIC2 31 STS 74 237 22Sf 22S2 143 22S4 2255 69 1 + + + + + + + + + + + 2 - + + + + + + + + + + 3 + + + - + + + + + + + 4 + + - - + + + + + + + 5 + + - - + + + + + + + 6 - - - - - + + + + + + 7 + + - - - + + + + + + 8 ------+ + + + + 9 ------+ + + + + 10 + + - - - - + + + + + 11 ------+ + + + + 12 + + - - - - + + + + + 13 + + + + + + + 14 + + - - - - - + + + + 15 ------+ + + 16 ------+ + + + and - indicate presence or absence of hybridization. For the pseudoautosomal locus MIC2, + and - indicate double and single dosage, respectively. Patient 1 is an exception, since he shows a triple dosage for MIC2. For loci DXS31, D YS74, STS, and DXS69, which share homology with the Y long arm, + and - indicate presence or absence of X-specific restriction bands. Interval k spans from the proximal breakpoint ofpatient 14 cent genes, such as adrenal hypoplasia, glycerol kinase, or to the proximal breakpoints ofpatients 15 and 16 and contains chronic granulomatous disease genes, causing contiguous the DXS143 locus. gene syndromes (31, 32). The study of patients with such Interval I is defined only by the proximal breakpoints of deletions has represented the key to the mapping and cloning patients 15 and 16, and contains DNF22S4, DNF22S5, and of the DMD gene (33). On the long arm of the X chromosome DXS69 loci. deletions spanning the q21 locus may cause complex syndromes characterized by choroideremia, mental retardation, and deafness (34). Recently a contiguous gene syndrome in DISCUSSION the Miller-Dieker region, on the short arm of chromosome The term "contiguous gene syndromes" has been used to 17, has been described and characterized (35). describe a number of complex inherited syndromes due to We demonstrated here that contiguous gene syndromes deletions of adjacent disease genes (29, 30). In the p21 region also occur in the distal short arm of the X chromosome. The of the X chromosome deletions involving the Duchenne following complex syndromes have been observed in our muscular dystrophy (DMD) gene sometimes extend to adja- patients: SS + CDPX + MRX + XLI, SS + CDPX + MRX

.3 Xp22 .2 MIC2 .1 CDPX

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 FIG. 1. Diagram of Xp22-pter deletions. Deletions have been ordered from left to right according to the proximal breakpoint. Deletions intervals are indicated on the right by letters a to 1. The assignment of genes and disease loci (bold type) and anonymous sequences (plain type) to specific intervals is also shown. *Order of intervals e, f, and g could not be defined. Downloaded by guest on September 27, 2021 10004 Genetics: Ballabio et al. Proc. Natl. Acad. Sci. USA 86 (1989) 1 2 3 4 5 6 7 8 9 (patient 2). Our results clearly map CDPX distal to DXS3J. Since genetic evidence suggests that the gene for recessive CDPX is not pseudoautosomal, we tentatively localize CDPX ,; .9. I"- e -*- proximal to MIC2. iq.- MRX. It has been suggested previously that deletions or (I --w -S- c -*-Wi mutations of X chromosome loci may cause MRX (37). X-linked mental retardation occurs in approximately 1 in 600

. N, male births and fragile X syndrome accounts for only one- quarter of them (38). We found a correlation between a deletion of a specific region of the X chromosome and the presence of mental retardation as an additional sign in the patient. Patients with terminal deletions of Xp22.3-pter involving both DXS31 and 1) -- STS always show mild mental retardation. Two patients (cases 12 and 13) with XLI + KAL did not show mental retardation. These results map STS as a proximal limit for the mapping ofMRX. Furthermore, we studied two patients with an interstitial deletion in Xp22.3 showing MRX (cases 4 and 14) and a patient with a terminal deletion showing normal intelligence (case 2). Since DXS31 was retained in each case, these results map DXS31 as a distal limit. Therefore, we tentatively assigned a gene for MRX to Xp22.3, proximal to DXS31 and distal to STS. This may represent the gene (MRX2) previously mapped by Arveiler et al. (37) in a linkage study performed in one informative family. However, in that study MRX2 was mapped proximal to both DXS31 and FIG. 2. DNA analysis of seven patients with deletions in Xp22. DXS143 and, therefore, proximal to STS. This discrepancy The restriction enzyme used was EcoRI and the probe was G1.3. may be due to the incomplete penetrance ofthe disease in that Lane 1, male control; lanes 2-6, patients with deletion of the STS family or to the presence of two or more different loci for gene showing isolated XLI (lanes 2 and 5 correspond to patients 7 and MRX in the Xp22 region. 4, respectively); lanes 7 and 8, patients with deletion of the STS gene XLI/STS Deficiency. We have shown that patients with showing XLI associated with KAL (cases 14 and 12, respectively); isolated XLI/STS deficiency (cases 3, 4, 5, and 7, and 12 lane 9: female control. a, locus DNF22SJ; b, DNF22S2; c, DNF22S3; unrelated patients indicated as "case 10") have heterogeneous d, DNF22S4; e, DNF22S5. Locus DNF22SJ is deleted in patients deletion breakpoints. STS can be assigned to two corresponding to lanes 3, 4, 6, 7, and 8, whereas locus DNF22S2 is different deleted only in the patient corresponding to lane 7 (case 14), affected deletion intervals (e and f) separated by the breakpoint of by XLI + KAL. patient 3, a patient with a partial deletion of the STS gene (10). KAL. KAL has been previously localized proximal to the + XLI + KAL, MRX + XLI + KAL, MRX + XLI, and XLI STS locus on the basis of two findings: (i) the description of + KAL. DNA analysis allowed us to identify 12 deletion two families where KAL and XLI cosegregated (16, 17), and intervals and to map more precisely the following disease (ii) the absence of symptoms related to KAL in individuals loci: affected by XLI and carrying terminal deletions of Xp (15). SS. Both environmental factors and genetic background Previous molecular analyses, however, failed to detect differ- may contribute to the development of normal stature. It has ences in the extent ofdeletions between patients with isolated been proposed that a locus affecting height might be located XLI and patients with XLI associated with KAL (10). on the distal portion ofXp and Yp Patient 1 has the XXY Here we demonstrate that locus DNF22SJ is deleted in all (20). + chromosome constitution of the asso- patients with XLI KAL and is also deleted in 11 out of 14 Klinefelter syndrome, patients ciated with an X/Y translocation and a presumed deletion in with isolated XLI. This maps DNF22SJ between the pseudoautosomal in both STS and KAL. Furthermore, locus DNF22S2 is retained in all region Xp and Yp. Klinefelter patients with isolated XLI, whereas it is deleted in 3 out of 5 syndrome is generally associated with an increase in height; patients with XLI associated with KAL. however, patient 1 is 155 cm tall. Furthermore, both affected Therefore, we demonstrate that KAL maps to Xp22.3 males and heterozygous females with terminal deletions of (interval i) and is flanked by DNF22SJ on the distal side and Xp display SS. Therefore, we tentatively assign a gene by DNF22S2 on the proximal side. affecting height to the pseudoautosomal region, distal to the The Xg blood group has not been included in our study MIC2 locus (interval a). because in most cases the analysis was not informative. CDPX. Two forms of CDPX have been described. The However, it has been reported that Xg was not deleted in a dominant form has been reported exclusively in females and family with XLI + KAL (16). This result, together with the is presumably lethal in hemizygous males. The recessive observation that Xg was deleted in a patient with a terminal form is clinically milder and has been mapped to Xp22.3-pter deletion of Xp22.3-pter (13), maps Xg distal to STS. on the basis of different reports of patients with X/Y trans- Terminal deletions have been defined so by both cytoge- locations or carrying terminal deletions of Xp (14, 15, 18, 19, netic and molecular studies; however, since MIC2 was the and 36). Most of these cases were not characterized by most distal locus studied, the presence of X-telomeric se- molecular methods and in some ofthem STS activity was not quences in these cases cannot be ruled out. investigated. The description of two families with an inter- We observed two patients (cases 15 and 16) with deletions stitial deletion involving both STS and KAL (16, 17) and the involving intervals a to k. This demonstrates that the absence fact that none of the individuals carrying X/Y translocations ofthis region ofthe X chromosome is still compatible with life analyzed was apparently affected by KAL suggested the in the affected males. order Xpter-CDPX-XLI-KAL-cen. Our results have both clinical and genetic interest. We now report the molecular characterization of a patient On the basis of our observations, patients diagnosed as carrying an X/Y translocation affected by isolated CDPX affected with XLI, CDPX, or KAL should receive careful Downloaded by guest on September 27, 2021 Genetics: Ballabio et al. Proc. Natl. Acad. Sci. USA 86 (1989) 10005 clinical evaluation to detect the presence of a contiguous gene 6. Koenig, M., Camerino, G., Heilig, R. & Mandel, J. L. (1984) Nucleic Acids Res. 12, 4097-4109. syndrome. Some features of CDPX are age dependent, there- 7. Bickmore, W. A. & Cooke, H. J. (1987) Nucleic Acids Res. 15, 6261- fore diagnosis may be difficult in older patients. KAL may be 6271. overlooked because hypogonadism is less evident in younger 8. Yen, P. H., Allen, E., Marsh, B., Mohandas, T., Wang, N., Taggart, and tests are not R. T. & Shapiro, L. J. (1987) Cell 49, 443-454. patients olfactory routinely performed in all 9. Fraser, N., Ballabio, A., Zollo, M., Persico, M. G. & Craig, 1. (1987) patients unless specifically requested. In case 11 the diagnosis Development 101, Suppl, 127-132. of KAL was made 8 years after the first examination, and in 10. Ballabio, A., Carrozzo, R., Parenti, G., Gil, A., Zollo, M., Persico, case 16 we were able to predict, on the basis ofmolecular data, M. G., Gillard, E., Affara, N., Yates, J., Ferguson-Smith, M. A., Frants, R. R., Eriksson, A. W. & Andria, G. (1989) Genomics 4, 36-40. that the patient was affected by KAL. This was confirmed 11. Ballabio, A., Carrozzo, R., Gil, A., Gillard, B., Affara, N., Ferguson- when the patient was subsequently reexamined. Smith, M. A., Fraser, N., Craig, I., Rocchi, M., Romeo, G. & Andria, Both cytogenetic and molecular analysis should be per- G. (1989) Ann. Hum. Genet. 53, 9-14. 12. Tiepolo, L., Zuffardi, O., Fraccaro, M., di Natale, D., Gargantini, L., formed to screen for the presence of a deletion in Xp22-pter. Muller, C. R. & Ropers, H. H. (1980) Hum. Genet. 54, 205-206. An early diagnosis may have important implications for 13. Ferguson-Smith, M. A., Sanger, R., Tippett, P., Aitken, D. A. & Boyd, treatment, particularly in patients with KAL. Carrier detec- E. (1983) Cytogenet. Cell Genet. 32, 273-274. 14. Curry, C. J. R., Lanman, J. T., Tasai, J., O'Lague, P., Magenis, R. E., tion and prenatal diagnosis are feasible in some families. Brown, M., Goodfellow, P., Mohandas, T., Bergner, E. A. & Shapiro, Neither the pathogenetic mechanism nor the molecular L. J. (1984) N. Engl. J. Med. 311, 1010-1014. defect have been elucidated in CDPX and KAL. It has been 15. Ballabio, A., Parenti, G., Carrozzo, R., Coppa, G., Felici, L., Migliori, V., Silengo, M., Franceschini, P. & Andria, G. (1988) Clin. Genet. 34, suggested that CDPX may share a common pathogenetic 31-37. mechanism with Warfarin embryopathy and vitamin K ep- 16. Ballabio, A., Parenti, G., Tippett, P., Mondello, C., Di Maio, S., Tenore, oxide reductase deficiency since they both display similar A. & Andria, G. (1986) Hum. Genet. 72, 237-240. manifestations (19). KAL has been considered a true multiple 17. Andria, G., Ballabio, A., Parenti, G., Di Maio, S. & Piccirillo, A. (1984) J. Inherited Metab. Dis. 7, Suppl. 2, 158-160. congenital anomaly (39). At least two developmental field 18. Sunohara, N., Sakuragawa, N., Satoyoshi, E., Tanae, A. & Shapiro, defects have been observed in patients with KAL, the first L. J. (1986) Ann. Neurol. 19, 174-181. involving the central nervous system, including hypothala- 19. Bick, D., Curry, C. J. R., McGill, J. R., Schorderet, D. F., Bux, R. C. mus and olfactory bulbs and tracts, and the second & Moore, C. M. (1989) Am. J. Med. Genet. 33, 100-107. involving 20. Zuffardi, O., Maraschio, P., Lo Curto, F., Muller, U., Giarola, A. & the urogenital system. Recent work in central nervous system Perotti, L. (1982) Am. J. Med. Genet. 12, 175-184. development may explain the brain malformations seen in 21. Camerino, G., Grzeschik, K. H., Jaye, M., De La Salle, H., Tolstoshev, KAL (40, 41). P., Lecocq, J. P., Heilig, R. & Mandel, J. L. (1984) Proc. Natl. Acad. Some described in this be used to search Sci. USA 81, 498-502. probes study may 22. Mondello, C., Ropers, H. H., Craig, I. W., Tolley, E. & Goodfellow, for deletion in patients with CDPX and KAL. It will be of P. N. (1987) Ann. Hum. Genet. 51, 137-143. interest to establish if these disorders, as isolated entities 23. Affara, N. A., Ferguson-Smith, M., Magenis, R., Tolmie, J., Boyd, E., (i.e., not included in a contiguous gene syndrome), are also Cooke, A., Jamieson, D., Kwok, K., Mitchell, M. & Snadden, L. (1987) due to deletions. In a recent study on 57 with STS Nucleic Acids Res. 15, 7325-7342. patients 24. Gillard, E., Affara, N. A., Yates, J., Goudie, D. R., Lambert, J., Aitken, deficiency we reported that 84% of them display a deletion D. A. & Ferguson-Smith, M. A. (1987) Nucleic Acids Res. 15, 3977-3985. (10). This leads us to speculate that the same may also apply 25. Bardoni, B., Guioli, S., Raimondi, E., Heilig, R., Mandel, J. L., Otto- to other diseases whose genes are located nearby. lenghi, S. & Camerino, G. (1988) Genomics 3, 32-38. DFN22SJ and DFN22S2 loci have been demonstrated to 26- Middlesworth, W., Bertelson, C., Kunkel, L. M. (1985) Nucleic Acids Res. 13, 5723. flank the KAL gene on each side. These loci represent the 27. Kunkel, L. M., Tantravahi, U., Kurnit, D. M., Eisenhardt, M., Bruns, closest markers to this gene and may be used as starting points G. P. & Latt, S. A. (1983) Nucleic Acids Res. 11, 7961-7979. for chromosome walking or jumping toward the KAL gene. 28. Ballabio, A., Parenti, G., Carrozzo, R., Sebastio, G., Andria, G., Buckle, A deletion map ofthe distal short arm ofthe X chromosome V., Fraser, N., Boyd, Y., Craig, I., Rocchi, M., Romeo, G., Jobsis, A. C. & Persico, M. G. (1987) Proc. Natl. Acad. Sci. USA 84, 4519-4523. provides useful information for any attempt at regional clon- 29. Schmickel, R. D. (1986) J. Pediatr. 109, 231-241. ing. The resulting deletion panel may also be used for fine 30. Emanuel, B. S. (1988) Am. J. Hum. Genet. 43, 575-578. mapping of any newly cloned sequence in this region. 31. Francke, U., Ochs, H. D., de Martinville, B., Giacalone, J., Lindgren, V., Disteche, C., Pagon, R. A., Hofker, M. H., van Ommen, G. J. B., Note Added in Proof. X-linked ocular albinism (OA1) has been Pearson, P. & Wedgwood, R. J. (1985) Am. J. Hum. Genet. 37, 250-267. reported as cosegregating with XLI in one family (42). No apparent 32. McCabe, E. R. B., Towbin, J., Chamberlain, J., Baumbach, L., Witko- signs of OA1 have been observed in any of the 27 patients analyzed ski, J., van Ommen, G. J. B. & Koenig, M. (1989) J. Clin. Invest. 83, in this study. Specific ophthalmological evaluation ruled out the 95-99. presence of OA1 in patient 15, thus suggesting that the OA1 locus is 33. Kunkel, L. M., Monaco, A. P., Middlesworth, W., Ochs, H. & Latt, S. (1985) Proc. Natl. Acad. Sci. USA 82, 4778-4782. proximal to deletion interval k in our map. 34. Cremers, F. P. M., van de Pol, D. J. R., Diergaarde, P. J., Wieringa, B., We thank Dr. P. Goodfellow for probe p2B, Dr. L. Kunkel for Nussbaum, R. L., Schwartz, M. & Ropers, H. H. (1989) Genomics 4, dic56 and and Dr. J. L. Mandel for We 41-46. probes 71-7A, probe M1A. 35. Ledbetter, D. H., Ledbetter, S. A., vanTuinen, P., Summers, K. M., also thank Dr. C. J. R. Curry for clinical information and a cell line Robinson, T. J., Nakamura, Y., Wolff, R., White, R., Barker, D. F., of patient 16. We are indebted to Dr. D. Ledbetter for valuable Wallace, M. R., Collins, F. S. & Dobyns, W. B. (1989) Proc. Natl. Acad. suggestions and to Dr. C. T. Caskey, Dr. T. Webster, Dr. J. Sci. USA 86, 5136-5140. Chamberlain, and Dr. G. Persico for critical reading of the manu- 36. Agematsu, K., Koike, K., Morosawa, H., Nakahori, Y., Nakagome, Y. script. This work was supported by the "Progetto Strategico Genoma & Akabane, T. (1988) Hum. Genet. 80, 105-107. Umano" of the Consiglio Nazionale Ricerche to G.C. and by Grant 37. Arveiler, B., Alembik, Y., Hanauer, A., Jacobs, P., Tranebjaerg, L., SCE-0140 from the Commission of European Communities. Mikkelsen, M., Puissant, H., Larget Piet, L. & Mandel, J. L. (1988) Am. J. Med. Genet. 30, 473-483. 1. Shapiro, L. J. (1985) Adv. Hum. Genet. 14, 331-389. 38. Suthers, G. K., Turner, G. & Mulley, J. C. (1988) Am. J. Med. Genet. 2. Goodfellow, P., Banting, G., Sheer, D., Ropers, H. H., Caine, A., 30, 485-491. Ferguson-Smith, M. A., Povey, S. & Voss, R. (1983) Nature (London) 39. Wegenke, J. D., Uehling, D. T., Wear, J. B., Gordon, E. S., Bargman, 302, 346-349. J. G., Deacon, J. S. R., Herrmann, J. P. R. & Opitz, J. M. (1975) Clin. 3. Cooke, H. J., Brown, W. R. A. & Rappold, G. A. (1985) Nature (Lon- Genet. 7, 368-381. don) 317, 687-697. 40. Brunjes, P. C. & Frazier, L. L. (1986) Brain Res. Rev. 11, 1-45. 4. Rouyer, F., Simmler, M. C., Johnsson, C., Vergnaud, G., Cooke, H. J. 41. Schwanzel-Fukuda, M. & Pfaff, D. W. (1989) Nature (London) 338, & Weissenbach, J. (1986) Nature (London) 319, 291-295. 161-164. 5. Page, D. C., Mosher, R., Simpson, E. M., Fisher, E. M C. Mardon, 42. Schnur, R. E., Punnett, H. H., Kistenmacher, M., Naids, R., Tomeo, G., Pollack, J., McGillivray, B., de la Chapelle, A. & Brown, L. G. 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