Chromosomal Translocations Mediated by Palindromic DNA

There is evidence accumulating to suggest that non-B DNA structures have a potential for genomic instability that induces genomic rearrangements including translocations and deletions. One of the best studied examples is the recurrent t(11;22) constitutional translocation in humans that is mediated by palindromic AT-rich repeats (PATRRs) on chromosomes 11q23 and 22q11. Cloned breakpoint sequences favor adopting a cruciform configuration in vitro. Analysis of the junction fragments implicates frequent double-strand-breaks at the center of both palindromic regions, followed by repair through the non-homologous end joining pathway. De novo examples of the translocation are detected at a substantial frequency in sperm samples from normal healthy males, but not in other normal somatic tissues or cell lines derived from human. Further our recent findings indicate that polymorphism of the PATRR affects the frequency of de novo translocation events and symmetrical alleles preferentially generate the translocation. We propose that the symmetric PATRR is likely to adopt a cruciform structure in male meiotic cells, creating genomic instability that leads to the recurrent translocation.     

Palindrome-mediated chromosomal translocations in humans

Recently, it has emerged that palindrome-mediated genomic instability contributes to a diverse group of genomic rearrangements including translocations, deletions, and amplifications. One of the best studied examples is the recurrent t(11;22) constitutional translocation in humans that has been well documented to be mediated by palindromic AT-rich repeats (PATRRs) on chromosomes 11q23 and 22q11. De novo examples of the translocation are detected at a high frequency in sperm samples from normal healthy males, but not in lymphoblasts or fibroblasts. Cloned breakpoint sequences preferentially form a cruciform configuration in vitro. Analysis of the junction fragments implicates frequent double-strand-breaks (DSBs) at the center of both palindromic regions, followed by repair through the non-homologous end joining (NHEJ) pathway. We propose that the PATRR adopts a cruciform structure in male meiotic cells, creating genomic instability that leads to the recurrent translocation.     

Fine structure DNA mapping studies of the chromosomal region harboring the genetic defect in neurofibromatosis type 1

To better map the location of the von Recklinghausen neurofibromatosis (NF1) gene, we have characterized a somatic cell hybrid designated 7AE-11. This microcell-mediated, chromosome-transfer construct harbors a centromeric segment and a neo-marked segment from the distal long arm of human chromosome 17. We have identified 269 cosmid clones with human sequences from a 7AE-11 library and, using a panel of somatic cell hybrids with a total of six chromosome 17q breakpoints, have mapped 240 of these clones on chromosome 17q. The panel included a hybrid (NF13) carrying a der(22) chromosome that was isolated from an NF1 patient with a balanced translocation, t(17;22) (q11.2;q11.2). Fifty-three of the cosmids map into a region spanning the NF13 breakpoint, as defined by the two closest flanking breakpoints (17q11.2 and 17q11.2-q12). RFLP clones from a subset of these cosmids have been mapped by linkage analysis in normal reference families, to localize the NF1 gene more precisely and to enhance the potential for genetic diagnosis of this disorder. The cosmids in the NF1 region will be an important resource for testing DNA blots of large-fragment restriction-enzyme digests from NF1 patient cell lines, to detect rearrangements in patients' DNA and to identify the 17;22 NF1 translocation breakpoint.     

A Palindrome-Mediated Recurrent Translocation with 3:1 Meiotic Nondisjunction: The t(8;22)(q24.13;q11.21)

Palindrome-mediated genomic instability has been associated with chromosomal translocations, including the recurrent t(11;22)(q23;q11). We report a syndrome characterized by extremity anomalies, mild dysmorphia, and intellectual impairment caused by 3:1 meiotic segregation of a previously unrecognized recurrent palindrome-mediated rearrangement, the t(8;22)(q24.13;q11.21). There are at least ten prior reports of this translocation, and nearly identical PATRR8 and PATRR22 breakpoints were validated in several of these published cases. PCR analysis of sperm DNA from healthy males indicates that the t(8;22) arises de novo during gametogenesis in some, but not all, individuals. Furthermore, demonstration that de novo PATRR8-to-PATRR11 translocations occur in sperm suggests that palindrome-mediated translocation is a universal mechanism producing chromosomal rearrangements.     

Precise localization of NF1 to 17q11.2 by balanced translocation.

A female patient is described with von Recklinghausen neurofibromatosis (NF1) in association with a balanced translocation between chromosome 17 and 22 [46,XX,t(17;22)(q11.2;q11.2)]. The breakpoint in chromosome 17 is cytogenetically identical to a previously reported case of NF1 associated with a 1;17 balanced translocation and suggests that the translocation events disrupt the NF1 gene. This precisely maps the NF1 gene to 17q11.2 and provides a physical reference point for strategies to clone the breakpoint and therefore the NF1 gene. A human-mouse somatic cell hybrid was constructed from patient lymphoblasts which retained the derivative chromosome 22 (22pter----22q11.2::17q11.2----17qter) but not the derivative 17q or normal 17. Southern blot analysis with genes and anonymous probes known to be in proximal 17q showed ErbA1, ErbB2, and granulocyte colony-stimulating factor (CSF3) to be present in the hybrid and therefore distal to the breakpoint, while pHHH202 (D17S33) and beta crystallin (CRYB1) were absent in the hybrid and therefore proximal to the breakpoint. The gene cluster including ErbA1 is known to be flanked by the constitutional 15;17 translocation breakpoint in hybrid SP3 and by the acute promyelocytic leukemia (APL) breakpoint, which provides the following gene and breakpoint order: cen-SP3-(D17S33,CRYB1)-NF1-(CSF3,ERBA1, ERBB2)-APL-tel. The flanking breakpoints of SP3 and API are therefore useful for rapidly localizing new markers to the neurofibromatosis critical region, while the breakpoints of the two translocation patients provide unique opportunities for reverse genetic strategies to clone the NF1 gene.     

Cruciform extrusion propensity of human translocation-mediating palindromic AT-rich repeats

There is an emerging consensus that secondary structures of DNA have the potential for genomic instability. Palindromic AT-rich repeats (PATRRs) are a characteristic sequence identified at each breakpoint of the recurrent constitutional t(11;22) and t(17;22) translocations in humans, named PATRR22 (~600bp), PATRR11 (~450bp) and PATRR17 (~190bp). The secondary structure-forming propensity in vitro and the instability in vivo have been experimentally evaluated for various PATRRs that differ regarding their size and symmetry. At physiological ionic strength, a cruciform structure is most frequently observed for the symmetric PATRR22, less often for the symmetric PATRR11, but not for the other PATRRs. In wild-type E. coli, only these two PATRRs undergo extensive instability, consistent with the relatively high incidence of the t(11;22) in humans. The resultant deletions are putatively mediated by central cleavage by the structure-specific endonuclease SbcCD, indicating the possibility of a cruciform conformation in vivo. Insertion of a short spacer at the centre of the PATRR22 greatly reduces both its cruciform extrusion in vitro and instability in vivo. Taken together, cruciform extrusion propensity depends on the length and central symmetry of the PATRR, and is likely to determine the instability that leads to recurrent translocations in humans.     

Comparative mapping of the constitutional and tumor-associated 11;22 translocations.

The reciprocal t(11;22)(q23;q11) is the most common non-Robertsonian constitutional translocation in humans. The tumor-associated 11;22 rearrangement of Ewing sarcoma (ES) and peripheral neuroepithelioma (NE) is cytologically very similar to the 11;22 constitutional rearrangement. Using immunoglobulin light-chain constant region, ETS1 probes, and the technique of in situ hybridization, we previously were able to show that the constitutional and ES/NE breakpoints are different. As a first step toward isolating these translocation junctions and to further distinguish between them, we have made somatic cell hybrids. Cells from a constitutional 46,XX,inv(9),t(11;22) carrier and from an ES cell line with a t(11;22) were separately fused to a hypoxanthine-guanine phosphoribosyltransferase-deficient Chinese hamster cell line (RJK88). Resulting clones were screened with G-banding and Southern hybridization. Hybrid clones derived from the constitutional t(11;22) were established which contained the der(22) and both the der(22) and the der(11). Hybrid clones derived from the ES cell line containing the der(11) were isolated. Using the technique of Southern hybridization we have sublocalized the loci; ApoA1/C3, CD3D, ETS1, PBGD, THY1, D11S29, D11S34, and D11S147 to the region between the two breakpoints on chromosome 11 and V lambda I, V lambda VI, V lambda VII, and D22S10 to the region between the breakpoints on chromosome 22. Using anonymous DNA probes, we found that D22S9 and D22S24 map proximal to the constitutional breakpoint and that D22S15 and D22S32 map distal to the ES breakpoint on chromosome 22.     

Tightly clustered 11q23 and 22q11 breakpoints permit PCR-based detection of the recurrent constitutional t(11;22)

Palindromic AT-rich repeats (PATRRs) on chromosomes 11q23 and 22q11 at the constitutional t(11;22) breakpoint are predicted to induce genomic instability, which mediates the translocation. A PCR-based translocation-detection system for the t(11;22) has been developed with PCR primers flanking the PATRRs of both chromosomes, to examine the involvement of the PATRRs in the recurrent rearrangement. Forty unrelated carriers of the t(11;22) balanced translocation, plus two additional, independent cases with the supernumerary-der(22) syndrome, were analyzed to compare their translocation breakpoints. Similar translocation-specific junction fragments were obtained from both derivative chromosomes in all 40 carriers of the t(11;22) balanced translocation and from the der(22) in both of the offspring with unbalanced supernumerary-der(22) syndrome, suggesting that the breakpoints in all cases localize within these PATRRs and that the translocation is generated by a similar mechanism. This PCR strategy provides a convenient technique for rapid diagnosis of the translocation, indicating its utility for prenatal and preimplantation diagnosis in families including carriers of the balanced translocation.     

Clustered 11q23 and 22q11 breakpoints and 3:1 meiotic malsegregation in multiple unrelated t(11;22) families

The t(11;22) is the only known recurrent, non-Robertsonian constitutional translocation. We have analyzed t(11;22) balanced-translocation carriers from multiple unrelated families by FISH, to localize the t(11;22) breakpoints on both chromosome 11 and chromosome 22. In 23 unrelated balanced-translocation carriers, the breakpoint was localized within a 400-kb interval between D22S788 (N41) and ZNF74, on 22q11. Also, 13 of these 23 carriers were tested with probes from chromosome 11, and, in each, the breakpoint was localized between D11S1340 and APOA1, on 11q23, to a region 相似文献   

Physical mapping of the von Recklinghausen neurofibromatosis region on chromosome 17

The von Recklinghausen neurofibromatosis (NF1) locus has been linked to chromosome 17, and recent linkage analyses place the gene on the proximal long arm. NF1 probably resides in 17q11.2, since two unrelated NF1 patients have been identified who possess constitutional reciprocal translocations involving 17q11.2 with chromosomes 1 and 22. We have used a somatic-cell hybrid from the t(17;22) individual, along with other hybrid cell lines, to order probes around the NF1 locus. An additional probe, 17L1, has been isolated from a NotI linking library made from flow-sorted chromosome 17 material and has been mapped to a region immediately proximal to the translocation breakpoint. While neither NF1 translocation breakpoint has yet been identified by pulse-field gel analysis, an overlap between two probes, EW206 and EW207, has been detected. Furthermore, we have identified the breakpoint in a non-NF1 translocation, SP-3, on the proximal side of the NF1 locus. This breakpoint has been helpful in creating a 1,000-kb pulsed-field map, which includes the closely linked NF1 probes HHH202 and TH17.19. The combined somatic-cell hybrid and pulsed-field gel analysis we report here favors the probe order D17Z1-HHH202-TH17.19-CRYB1-17L1-NF1- (EW206, EW207, EW203, L581, L946)-(ERBB2, ERBA1). The agreement in probe ordering between linkage analysis and physical mapping is excellent, and the availability of translocation breakpoints in NF1 should now greatly assist the cloning of this locus.     

Breakpoint mapping and haplotype analysis of three reciprocal translocations identify a novel recurrent translocation in two unrelated families: t(4;11)(p16.2;p15.4)

The majority of constitutional reciprocal translocations appear to be unique rearrangements arising from independent events. However, a small number of translocations are recurrent, most significantly the t(11;22)(q23;q11). Among large series of translocations there may be multiple independently ascertained cases with the same cytogenetic breakpoints. Some of these could represent additional recurrent rearrangements, alternatively they could be identical by descent (IBD) or have subtly different breakpoints when examined under higher resolution. We have used molecular breakpoint mapping and haplotyping to determine the origin of three pairs of reciprocal constitutional translocations, each with the same cytogenetic breakpoints. FISH mapping showed one pair to have different breakpoints and thus to be distinct rearrangements. Another pair of translocations were IBD with identical breakpoint intervals and highly conserved haplotypes on the derived chromosomes. The third pair, t(4;11)(p16.2;p15.4), had the same breakpoint intervals by aCGH and fosmid mapping but had very different haplotypes, therefore they represent a novel recurrent translocation. Unlike the t(11;22)(q23;q11), the formation of the t(4;11)(p16.2;p15.4) may have involved segmental duplications and sequence homology at the breakpoints. Additional examples of recurrent translocations could be identified if the resources were available to study more translocations using the approaches described here. However, like the t(4;11)(p16.2;p15.4), such translocations are likely to be rare with the t(11;22) remaining the only common recurrent constitutional reciprocal translocation.     

Der(22) syndrome and velo-cardio-facial syndrome/DiGeorge syndrome share a 1.5-Mb region of overlap on chromosome 22q11

Derivative 22 (der[22]) syndrome is a rare disorder associated with multiple congenital anomalies, including profound mental retardation, preauricular skin tags or pits, and conotruncal heart defects. It can occur in offspring of carriers of the constitutional t(11;22)(q23;q11) translocation, owing to a 3:1 meiotic malsegregation event resulting in partial trisomy of chromosomes 11 and 22. The trisomic region on chromosome 22 overlaps the region hemizygously deleted in another congenital anomaly disorder, velo-cardio-facial syndrome/DiGeorge syndrome (VCFS/DGS). Most patients with VCFS/DGS have a similar 3-Mb deletion, whereas some have a nested distal deletion endpoint resulting in a 1.5-Mb deletion, and a few rare patients have unique deletions. To define the interval on 22q11 containing the t(11;22) breakpoint, haplotype analysis and FISH mapping were performed for five patients with der(22) syndrome. Analysis of all the patients was consistent with 3:1 meiotic malsegregation in the t(11;22) carrier parent. FISH-mapping studies showed that the t(11;22) breakpoint occurred in the same interval as the 1.5-Mb distal deletion breakpoint for VCFS. The deletion breakpoint of one VCFS patient with an unbalanced t(18;22) translocation also occurred in the same region. Hamster-human somatic hybrid cell lines from a patient with der(22) syndrome and a patient with VCFS showed that the breakpoints occurred in an interval containing low-copy repeats, distal to RANBP1 and proximal to ZNF74. The presence of low-copy repetitive sequences may confer susceptibility to chromosome rearrangements. A 1.5-Mb region of overlap on 22q11 in both syndromes suggests the presence of dosage-dependent genes in this interval.     

Chromosomal translocations and palindromic AT-rich repeats

Repetitive DNA sequences constitute 30% of the human genome, and are often sites of genomic rearrangement. Recently, it has been found that several constitutional translocations, especially those that involve chromosome 22, take place utilizing palindromic sequences on 22q11 and on the partner chromosome. Analysis of translocation junction fragments shows that the breakpoints of such palindrome-mediated translocations are localized at the center of palindromic AT-rich repeats (PATRRs). The presence of PATRRs at the breakpoints indicates a palindrome-mediated mechanism involved in the generation of these constitutional translocations. Identification of these PATRR-mediated translocations suggests a universal pathway for gross chromosomal rearrangement in the human genome. De novo occurrences of PATRR-mediated translocations can be detected by PCR in normal sperm samples but not somatic cells. Polymorphisms of various PATRRs influence their propensity for adopting a secondary structure, which in turn affects de novo translocation frequency. We propose that the PATRRs form an unstable secondary structure, which leads to double-strand breaks at the center of the PATRR. The double-strand breaks appear to be followed by a non-homologous end-joining repair pathway, ultimately leading to the translocations. This review considers recent findings concerning the mechanism of meiosis-specific, PATRR-mediated translocations.     

The second case of a t(17;22) in a family with neurofibromatosis type 1: sequence analysis of the breakpoint regions

A reciprocal t(17;22)(q11.2;q11.2) was found in a female patient with neurofibromatosis type 1 (NF1) and in her affected daughter. Sequence analysis of cloned junction fragments traversing the breakpoints allowed the identification of the structures involved in the rearrangement. Aberrant bands in Southern hybridizations of restriction enzyme-digested DNA of the patient pointed to the disruption of the NF1 gene in intron 31. Semispecific polymerase chain reaction analysis of the genomic DNA of the patient with the specific primer anchored at NF1 exon 31 was used to obtain the breakpoint-spanning fragment of the derivative chromosome 17. The intron 31 sequence turned out to be interrupted within a large irregular (AT) repeat. The chromosome 22-derived sequence of the der(17) junction fragment allowed us to identify cosmids of the corresponding region from a chromosome 22-specific cosmid library. With the support of the breakpoint-spanning cosmids, the chromosome 22 region upstream of the fragment carried by the der(17) was characterized. Primers deduced from the sequence of this upstream region were used in combination with a primer in NF1 intron 31 distal to the breakpoint on chromosome 17 to amplify the der(22) junction fragment. The structure of the junction sequences suggested that the translocation had arisen by unequal homologous recombination between (AT)-rich repeats on chromosome 22 and on chromosome 17 in intron 31 of the NF1 gene. However, our data support the assumption of additional rearrangements prior to, or in the course of, the recombination event, leading to a loss of the sequences between the involved (AT) repeats on chromosome 22. In the direct vicinity of these (AT) repeats, two members of a previously undescribed low-copy repetitive sequence have been found, copies of which are also present on human chromosome 13. Received: 27 August 1996 / Revised: 7 October 1996     

The breakpoint region of the most common isochromosome, i(17q), in human neoplasia is characterized by a complex genomic architecture with large, palindromic, low-copy repeats

Although a great deal of information has accumulated regarding the mechanisms underlying constitutional DNA rearrangements associated with inherited disorders, virtually nothing is known about the molecular processes involved in acquired neoplasia-associated chromosomal rearrangements. Isochromosome 17q, or "i(17q)," is one of the most common structural abnormalities observed in human neoplasms. We previously identified a breakpoint cluster region for i(17q) formation in 17p11.2 and hypothesized that genome architectural features could be responsible for this clustering. To address this hypothesis, we precisely mapped the i(17q) breakpoints in 11 patients with different hematologic malignancies and determined the genomic structure of the involved region. Our results reveal a complex genomic architecture in the i(17q) breakpoint cluster region, characterized by large ( approximately 38-49-kb), palindromic, low-copy repeats, strongly suggesting that somatic rearrangements are not random events but rather reflect susceptibilities due to the genomic structure.     

Fine mapping of the constitutional translocation t(11;22)(q23;q11)

Translocation t(11;22)(q23;q11) is the most common constitutional reciprocal translocation in man. Balanced carriers are phenotypically normal, except for decreased fertility, an increased spontaneous abortion rate and a possible predisposition to breast cancer in some families. Here, we report the high resolution mapping of the t(11;22)(q23;q11) breakpoint. We have localised the breakpoint, by using fluorescence in situ hybidisation (FISH) walking, to a region between D11S1340 and WI-8564 on chromosome 11, and D22S134 and D22S264 on chromosome 22. We report the isolation of a bacterial artificial chromosome (BAC) clone spanning the breakpoint in 11q23. We have narrowed down the breakpoint to an 80-kb sequenced region on chromosome 11 and FISH analysis has revealed a variation of the breakpoint position between patients. In 22q11, we have sequenced two BACs (BAC2280L11 and BAC41C4) apparently mapping to the region; these contain low copy repeats (LCRs). Southern blot analysis with probes from BAC2280L11 has revealed different patterns between normal controls and translocation carriers, indicating that sequences similar/identical to these probes flank the translocation breakpoint. The occurrence of LCRs has previously been associated with genomic instability and "unclonable" regions. Hence, the presence of such repeats renders standard translocation breakpoint cloning techniques ineffective. Thus, we have used high resolution fiber-FISH to study this region in normal and translocation cases by using probes from 22q11, LCRs and 11q23. We demonstrate that the LCR containing the gap in 22q11 is probably substantially larger than the previous estimates of 100 kb. Using fiber-FISH, we have localised the breakpoint in 22q11 to approximately 20-40 kb from the centromeric border of the LCR (i.e. the telomeric end of AC006547) and have confirmed the breakpoint position on 11q23.     

AT-rich palindromes mediate the constitutional t(11;22) translocation

The constitutional t(11;22) translocation is the only known recurrent non-Robertsonian translocation in humans. Offspring are susceptible to der(22) syndrome, a severe congenital anomaly disorder caused by 3&rcolon;1 meiotic nondisjunction events. We previously localized the t(11;22) translocation breakpoint to a region on 22q11 within a low-copy repeat termed "LCR22" and within an AT-rich repeat on 11q23. The LCR22s are implicated in mediating different rearrangements on 22q11, leading to velocardiofacial syndrome/DiGeorge syndrome and cat-eye syndrome by homologous recombination mechanisms. The LCR22s contain AT-rich repetitive sequences, suggesting that such repeats may mediate the t(11;22) translocation. To determine the molecular basis of the translocation, we cloned and sequenced the t(11;22) breakpoint in the derivative 11 and 22 chromosomes in 13 unrelated carriers, including two de novo cases and der(22) syndrome offspring. We found that, in all cases examined, the reciprocal exchange occurred between similar AT-rich repeats on both chromosomes 11q23 and 22q11. To understand the mechanism, we examined the sequence of the breakpoint intervals in the derivative chromosomes and compared this with the deduced normal chromosomal sequence. A palindromic AT-rich sequence with a near-perfect hairpin could form, by intrastrand base-pairing, on the parental chromosomes. The sequence of the breakpoint junction in both derivatives indicates that the exchange events occurred at the center of symmetry of the palindromes, and this resulted in small, overlapping staggered deletions in this region among the different carriers. On the basis of previous studies performed in diverse organisms, we hypothesize that double-strand breaks may occur in the center of the palindrome, the tip of the putative hairpin, leading to illegitimate recombination events between similar AT-rich sequences on chromosomes 11 and 22, resulting in deletions and loss of the palindrome, which then could stabilize the DNA structure.     

Molecular cloning of Xp11 breakpoints in two unrelated mentally retarded females with X;autosome translocations

Mental retardation is a very common and extremely heterogeneous disorder that affects about 3% of the human population. Its molecular basis is largely unknown, but many loci have been mapped to the X chromosome. We report on two mentally retarded females with X;autosome translocations and breakpoints in Xp11, viz., t(X;17)(p11;p13) and t(X;20)(p11;q13). (Fiber-) FISH analysis assigned the breakpoints to different subbands, Xp11.4 and Xp11.23, separated by approximately 8 Mb. High-resolution mapping of the X- chromosome breakpoints using Southern blot hybridization resulted in the isolation of breakpoint-spanning genomic subclones of 3 kb and 0. 5 kb. The Xp11.4 breakpoint is contained within a single copy sequence, whereas the Xp11.23 breakpoint sequence resembles an L1 repetitive element. Several expressed sequences map close to the breakpoints, but none was found to be inactivated. Therefore, mechanisms other than disruption of X-chromosome genes likely cause the phenotypes.     

A common breakpoint on 11q23 in carriers of the constitutional t(11;22) translocation

Structural chromosomal rearrangements occur commonly in the general population. Individuals that carry a balanced translocation are at risk of having unbalanced offspring; therefore, the frequency of translocations in couples with recurrent spontaneous abortions is higher than that in the general population. The constitutional t(11;22) translocation is the most common recurrent non-Robertsonian translocation in humans and may serve as a model to determine the mechanism that causes recurrent meiotic translocations. We previously localized the t(11;22) translocation breakpoint to a region on 22q11 within a low-copy repeat, termed "LCR22." To define the breakpoint on 11q23 and to ascertain whether this region shares homology with LCR22 sequences, we performed haplotype analysis on patients with der(22) syndrome. We found that the breakpoint on 11q23 occurred between two genetic markers, D11S1340 and APOC3-tetra, both being present within a single bacterial-artificial-chromosome clone. To determine whether the breakpoint occurred within the same region among a larger set of carriers, we performed FISH mapping studies. The breakpoints were all within the same clone, suggesting that this region may harbor sequences that are prone to breakage. We narrowed the breakpoint interval, in both derivative chromosomes from two unrelated carriers, to a 190-bp, AT-rich repeat, which indicates that this repeat may mediate recombination events on chromosome 11. Interestingly, the LCR22s harbor AT-rich repeats, suggesting that this sequence motif may mediate recombination events in nonhomologous chromosomes during meiosis.     

Subregional physical mapping of an αB-crystallin sequence and of a new expressed sequence D11S877E to human 11q

We report the regional assignment on Chromosome (Chr) 11q of two cDNA clones selected as sequences expressed in mature kidney and not expressed in Wilms' tumor. Clone T70 was identified as an B-crystallin sequence (CRYA2). CRYA2 has previously been mapped to 11q22.3–23.1 by in situ hybridization. Clone 6.2 represents a new gene expressed in adult and fetal kidney, pancreas, and liver. In order to map sequences corresponding to clone 6.2 and to physically define the boundaries of the localization of CRYA2, we used somatic cell hybrids carrying either different human chromosomes or Chr 11 segments and a cell line established from a patient with an interstitial deletion of region 11q14.3–q22.1. We showed that CRYA2 lies proximal to the 11q23.2 breakpoint defined by the constitutional t(11;22) and distal to the 11q22.1 breakpoint (between D11S388 and D11S35) of a constitutional interstitial deletion. This is in agreement with previous data obtained by in situ hybridization and provides proximal and distal physical benchmarks for this localization. Clone 6.2-related sequence (D11S877E) was assigned to region 11q23.2–q24.2 defined by the breakpoints of the constitutional t(11;22) and of the Ewing's sarcoma neuroepithelioma t(11;22).