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Human Molecular Genetics Pages 431-437  

Allele-specific late replication and fragility of the most active common fragile site, FRA3B
Introduction
Results
   FRA3B expression and DNA replication in the FHIT/FRA3B region
   Profile of replication timing in the FHIT/FRA3B region
Discussion
Materials And Methods
   Cell lines and reagents
   Visualization of late replication in the FRA3B region
   Flow cytometry and DNA isolation
   PCR analysis of replicated DNA
   Replication timing analysis
Acknowledgements
Abbreviations
References

Allele-specific late replication and fragility of the most active common fragile site, FRA3B

Allele-specific late replication and fragility of the most active common fragile site, FRA3B

Liang Wang, John Darling, Jin-San Zhang, Haojie Huang, Wanguo Liu and David I. Smith*

Division of Experimental Medicine, Department of Laboratory Medicine and Pathology, Mayo Clinic/Foundation, 200 First Street SW, Rochester, MN 55905, USA

Received September 11, 1998; Revised and Accepted November 30, 1998

FRA3B at 3p14.2 is the most active of the common fragile sites in the human genome and is expressed when cells are exposed to the DNA replication inhibitor, aphidicolin. Several lines of evidence suggest that fragile sites are regions of late replication. To elucidate the relationship between the timing of replication across the FRA3B region and its corresponding fragility, we labeled cells with 5-bromo-2[prime]-deoxyuridine (BrdU) and adopted an immunofluorescent procedure to visualize late replicating DNA (BrdU-substituted DNA) in metaphase chromosomes. We also chose 21 markers along the FRA3B region and analyzed the timing of replication using BrdU-labeled DNA from different stages of the cell cycle sorted by flow cytometry. Our results show that there are two distinct alleles that replicate at different stages in the cell cycle and that breaks/gaps preferentially occurred on the chromosome 3 with the late replication allele. These results provide direct evidence that allele-specific late replication is involved in the fragility of the most active common fragile site, FRA3B.

INTRODUCTION

FRA3B is the most highly inducible fragile site in the human genome (1). Deletions in the FRA3B region are observed in a number of different solid tumors (2-6). Ohta et al. (7) identified the fragile histidine triad (FHIT) gene from this region, and they identified alterations in this gene using nested RT-PCR analysis in a number of different tumor types (7-9). The human FHIT gene spans an estimated 1 Mb of DNA at chromosome 3p14.2 and encompasses the entire FRA3B region and a t(3;8) translocation breakpoint observed in a family with hereditary renal cell carcinoma (the hRCC breakpoint). In contrast to its large genomic size, the final processed transcript of this gene is only 1.1 kb and it encodes a 16.8 kDa protein with diadenosine triphosphate hydrolase activity (10).

The molecular basis for the fragility in the FRA3B region is not yet known. Previous studies (11,12) have suggested that the common fragile site FRA3B is distinct from the cloned rare fragile sites. First, no repeat motifs, such as trinucleotide or minisatellite repeats, characteristic of rare fragile sites have been identified. Second, genomic breakage and instability occur over a large genomic region (at least 500 kb). It has been hypothesized that gaps at fragile sites arise as a result of replication failure at chromosomal points that are unusually sensitive to interference during DNA synthesis (13). Using a fluorescence in situ hybridization (FISH)-based procedure to analyze replication timing, Le Beau et al. (14) reported that FRA3B sequences are late replicating and that replication was delayed further by aphidicolin, an inhibitor of DNA polymerases [alpha] and [delta] (15).

Although the function of the FHIT gene has not yet been determined, the gene can be used as a biomarker to analyze DNA replication around the FHIT/FRA3B region. We treated cell lines with aphidicolin and analyzed replication timing in the FHIT region using multiplex PCR analysis with human FHIT exon-specific primers as well as markers throughout the FRA3B region including several polymorphic markers. We also adopted an immunofluorescent procedure to visualize 5-bromo-2[prime]-deoxyuridine (BrdU)-substituted DNA in metaphase chromosomes. Our data show that replication timing in the FHIT/FRA3B region is asynchronous and that there are two distinct alleles that replicate at different times in the cell cycle. Breaks/gaps within the FRA3B region preferentially occurred on the chromosome with the late replicating (LR) allele. To our knowledge, this is the first report to demonstrate allele-specific late replication in this region.

RESULTS

FRA3B expression and DNA replication in the FHIT/FRA3B region

To visualize the replication pattern across the FRA3B region, we labeled the cells with BrdU for 3 h and then collected them for chromosome preparation. Indirect immunofluorescence was used for detection of BrdU-substituted DNA in metaphases. Since BrdU was incorporated into DNA in the last 3 h before the cells entered M phase, the detected BrdU should represent the LR region on chromosomes. Preliminary data from cell line GM03715 and two normal blood samples showed that LR signals were observed mostly on one of the two chromosomes 3 and that breaks/gaps in the FRA3B region occurred preferentially on the chromosome with LR signals. These data raise an important question, which is whether FRA3B expression occurs randomly between the chromosome 3 homologs or preferentially on one homolog. To answer this question, we chose another cell line (GM11428) with abnormalities on both chromosomes 3, which were easily differentiable as one had a partial deletion, del(3), and the other had a partial duplication, dup(3). We analyzed 660 metaphases from this cell line, comprising 278 metaphases from cells with aphidicolin induction and 382 metaphases from cells without aphidicolin induction. We first analyzed late replication and compared the results obtained between aphidicolin-induced cells and cells not exposed to aphidicolin. After aphidicolin induction, a total of 75 LR signals out of 278 metaphases were identified in the FRA3B region while 48 of 382 metaphases from untreated cells showed LR signals in this fragile site. The difference between the two groups is statistically significant (P1 = 0.0001, [chi]2 test) (Table 1), demonstrating a delay of replication at FRA3B after aphidicolin induction. Figure 1 shows some representative metaphases with LR and/or fragile site breakage on one chromosome 3 while the other chromosome 3 did not have a BrdU LR signal or a fragile site break.


Figure 1. Visualization of late replication in the FRA3B region. Cultured cells were treated with aphidicolin for 24 h and labeled by BrdU for 3 h before harvest for chromosome preparation. BrdU-labeled DNA in the metaphase chromosomes was detected by an immunofluorescent procedure (see Materials and Methods). The green/light blue spots represent the late replicating region (BrdU-substituted DNA) on the chromosomes. The metaphase spreads were obtained from cell lines GM11428 (A and B) and GM03715 (C and D), respectively. (A) and (C) show late replication of one chromosome 3 homolog without fragile site breakage in either homolog. (B) and (D) show late replication of one homolog with fragile site breakage. Two of the metaphases (A and D) also showed very strong signals (late replication) on the inactive chromosomes X. del(3), chromosome 3 with partial deletion; dup(3), chromosome 3 with duplication.

Table 1. Preferential breaks at chromosome 3 with the late replication allele
  Metaphases analyzed LR signals
del(3)/dup(3)		 Breaks
del(3)/dup(3)
Aphidicolin induction 278 75 (63/12) 33 (31/2)
Control (no induction) 382 48 (48/0) 8 (8/0)
LR, late replication; del(3), chromosome 3 with deletion; dup(3), chromosome 3 with duplication.


Since the cell line utilized has two distinct homologs, del(3) and dup(3), it was very easy to differentiate them. We next analyzed LR and breaks/gaps between the two chromosome 3 homologs. Interestingly, 84% (63/75) of LR signals were detected on the chromosome 3 with the partial deletion in aphidicolin-treated cells (Table 1). A similar result was also obtained in the cells without aphidicolin treatment (Table 1). The difference between the two distinguishable chromosomes 3 in this cell line was significant (P2 = 0.035, [chi]2 test). These results indicate that replication at FRA3B is asynchronous. One chromosome 3 contains an LR allele and the other contains an earlier replicating allele. More interestingly, the chromosome with the LR allele showed more breaks/gaps at FRA3B than its homolog, especially after aphidicolin induction (Table 1), suggesting that late replication is associated with fragility of the fragile site FRA3B.

Profile of replication timing in the FHIT/FRA3B region

BrdU-labeled human DNAs from six fractions of the cell cycle were isolated by co-immunoprecipitation with BrdU-labeled hamster DNA. To test the quality of the isolated DNA, we performed control experiments by analyzing control PCR products in the cell line GM03715. First, the hamster-specific PCRs from hypoxanthine-guanine phosphoribosyltransferase (HXGPRT) and transducin (TRANA) genes showed constant bands across the six fractions of cell cycle, suggesting that BrdU-labeled human DNAs were co-precipitated successfully (data not shown). Secondly, human-specific PCR primers derived from regions with known replication profiles showed late (DXS297 and DXS7857) and early replication (HPRT) as expected, further suggesting that the DNA did indeed represent different fractions in the cell cycle (Fig. 2A). We then used the BrdU-substituted DNAs as templates to analyze the replication timing of FHIT exons and other markers along the FHIT/FRA3B region. The results showed that replication was clearly asynchronous in at least six of seven FHIT exons, including exons 3, 4, 6, 7, 8 and 9 (Fig. 3A). For example, replication of exon 8 started at S1, stopped at S2, and restarted at S3. After aphidicolin induction, the replication was delayed but still remained asynchronous (Fig. 3B). To determine if this actually represented asynchronous replication between two alleles, we analyzed three polymorphic markers from this region (D3S4260, D3S1600 and D3S1313) against the BrdU-substituted DNA. The result of this analysis demonstrated that there are indeed two distinct alleles with independent replication timing in the cell line analyzed (Fig. 2B).


Figure 2. Allele-specific replication in the FHIT/FRA3B region. PCR products were labeled and separated on sequencing gels. (A) A control experiment was performed to ensure the high quality of isolated DNAs. The PCR products from known replication pattern markers produced early (HPRT) and late (DXS297 and DXS7857) replication signals as expected. Note: both of the late replication markers also showed early replication bands since the DNAs were isolated from female cells with only one late replicated chromosome X (inactivated). (B) Three polymorphic markers from the FHIT/FRA3B region displayed asynchronous replication of their two distinct alleles. The arrows demonstrate the two different replicating alleles for each polymorphic marker.


Figure 3. Replication timing profile of the FHIT gene. BrdU-labeled DNA from six different fractions of cell cycle were subjected to multiplex PCR by direct labeling with [[alpha]-32P]dCTP followed by separation on sequencing gels (see Materials and Methods). (A) Replication profile before aphidicolin induction. (B) Replication profile after aphidicolin induction. Asynchronous replication timing can be seen with most of the FHIT exons.

To better understand the replication profile in the FHIT/FRA3B region, we scanned all of the signals and compared the relative density of each signal. We chose the two cell cycle stages that gave the greatest amplification with each respective marker to represent the replication timing of the early and late alleles for that specific marker. Figure 4 includes all markers tested within the FHIT/FRA3B region and shows the replication pattern of the two respective alleles for each marker. Also included in this figure is the replication of the two alleles for each marker in the presence of aphidicolin. It can be clearly seen that most of the markers replicate later (for both alleles) in the presence of aphidicolin. The LR allele of FHIT, which usually replicates very late, replicates extremely late in the presence of aphidicolin: 15 of 21 markers tested showed replication of the LR allele in the G2/M phase in the presence of aphidicolin (Fig. 4). We also repeated this experiment in another cell line GM00546. Although the data are not shown here, the results from this cell line were similar to those from GM03715, suggesting that the allele-specific late replication within the FHIT/FRA3B region was not limited to one cell line.


Figure 4. Timing of the replication profile across the FHIT/FRA3B region in cell line GM03715 in the absence and presence of aphidicolin. A total of 21 markers across the FHIT/FRA3B region were analyzed. The approximate position of each marker within the FHIT/FRA3B region is shown on the left. Also included in this figure is the region where aphidicolin-inducible breakage occurs (the FRA3B region), as well as important landmarks in that region including the hRCC breakpoint, the proximal and distal clusters of aphidicolin-induced breakpoints, the position of the HPV16 viral integration site and the position of pSV2neo insertions into the FRA3B region in the presence of aphidicolin. The two strongest signals were chosen to represent the replication timing for each allele for that marker. The curved lines represent replication timing for each allele across this region. The most significant asynchronous replication between the two alleles occurred at FHIT exon 6, D3S1300, D3S4482, FHIT exon 5, U39804, U39799, spc-DNA1, FHIT exon 4, D3S1480 and FHIT exon 3 (arrows beside each marker).

DISCUSSION

Chromosome replication is under stringent temporal and spatial control to ensure accurate duplication of the genome. Timing of DNA replication appears to be an important epigenetic regulator of gene expression during development. Using a FISH-based procedure, Le Beau et al. (14) reported LR in the FRA3B region and this replication was delayed further in the presence of aphidicolin. By detecting BrdU-labeled DNA, we cytologically and molecularly demonstrate not only late replication but also allele-specific replication within the FHIT/FRA3B region, especially after aphidicolin induction (Table 1 and Fig. 4). After induction, the chromosomes with LR signals within the FRA3B region increased from 12.6 (48/382) to 27.0% (75/278), and chromosomes with breaks/gaps at the same region increased from 2.1 (8/382) to 11.9% (33/278) (Table 1). This demonstrates a coincidence between a delay of replication and breaks/gaps at FRA3B. More importantly, our data also show that the chromosome with the LR allele had more breaks/gaps than its homolog (Table 1), suggesting that indeed late replication and possibly incomplete replication may lead to breaks/gaps in the FHIT/FRA3B region. It is hypothesized that unreplicated DNA regions would affect the localized chromatin structure, and this might lead to the unstable as well as recombinogenic properties of this fragile site. The data herein provide direct evidence linking late replication and breaks/gaps to the common fragile site FRA3B.

Asynchronous DNA replication has been reported in imprinted regions such as 15q11-13, 11q23 and 11p15.5 (16-19). Imprinted regions appear to be unstable in the human genome since frequent deletions and rearrangements were observed in these regions in human cancers and also in several genetic diseases. So far, the molecular basis of the instability at imprinted regions is not yet known. Imprinted regions and the FRA3B region share several common features such as asynchronous replication and frequent deletion in cancers. Preliminary studies in our laboratory have shown that one allele of the FHIT gene in cancer cell lines is deleted preferentially (unpublished data) whereas the other allele appears to be unperturbed. This could explain why full-length RT-PCR products were still observed in cell lines with heterogeneous deletions in FHIT exons (20). Asynchronous replication may be associated with frequent splicing alterations within the FHIT gene, as seen in a variety of tumors and some normal tissues (21,22), since altered RNA splicing in other regions with asynchronous replication has been reported (23,24). More interestingly, FMR1 and FMR2, the two cloned genes at rare fragile sites, also demonstrated altered RNA splicing and late replication (25-28). Further analysis will help to address any relationship between DNA replication and RNA splicing.

The data presented here show that the markers throughout this region replicate at different times in the cell cycle (Fig. 4). This does not seem consistent with what we know about DNA replication, replicons and replication domains in other regions. This asynchronous replication pattern could be one of the important factors leading to fragility in the FRA3B region. Thus far, we are not sure what mechanism causes this unique replication pattern. Previous studies have shown that the DNA sequences in this region are high in A-T content, LINEs and MER repeats, whereas the number of Alu elements is reduced (12). Further studies are needed to determine what kind of DNA sequences or three-dimensional structures are associated with the asynchronous replication timing across this region.

In summary, we report here that there is allele-specific replication of the FHIT/FRA3B region. The expression of fragility in the FRA3B region preferentially occurred on the chromosome with the late replicating allele. The results herein provide evidence that allele-specific late replication is related to the fragility of the common fragile site FRA3B. It will be interesting to see if the genes that reside on the 87 other common fragile sites show allele-specific replication. Such an observation would suggest that this is the mechanism underlying the common fragile sites: a very late replicating allele, which cannot finish replication before cell division in the presence of aphidicolin or some other cellular stress, predisposes that homolog to breakage and rearrangement.

MATERIALS AND METHODS

Cell lines and reagents

Human lymphoblastoid cell lines GM03715, GM00546 and GM11428 were purchased from NIGMS. The former two cell lines are karyotypically normal. GM11428, came from a 6-year-old female with the following karyotype: 46, XX, del(3)(p25pter),der(3)add(3)(pter)dup(3)(q21qter). The two chromosomes 3 in this patient are easily distinguishable from each other, thus facilitating our analyses of chromosome-specific late replication. The culture conditions were based on recommendations from the supplier. In addition, two peripheral blood samples were obtained from two females with normal karyotypes. Aphidicolin and BrdU were purchased from Sigma (St Louis, MO). Mouse anti-BrdU antibody and fluorescein-labeled secondary antibody (anti-IgG) were obtained from Calbiochem (La Jolla, CA).

Visualization of late replication in the FRA3B region

Cell lines GM03715 and GM11428 and two whole blood samples were cultured in RPMI-1640 with 10% fetal bovine serum (FBS) and treated with aphidicolin (0.4 µM) for 24 h. The cells were then labeled with BrdU at a final concentration of 10 µM (for GM03715 and GM11428) or 50 µM (for the two whole blood samples) for 3 h and treated with colcemid at a final concentration of 0.4 µM for 1 h before harvesting. The cells were collected and subjected to chromosome preparation. Chromosomes from metaphase spreads were denatured by incubation for 2 min in a mixture of ethanol/0.1 M NaOH (2:5) (29). BrdU-labeled chromosomes were visualized by using an indirect immunofluorescence procedure. Briefly, the denatured slides were permeabilized with 0.1% NP-40 for 15 min followed by incubation with 10% goat serum in phosphate-buffered saline (PBS) (pH 7.4) for 20 min. The slides were incubated with 5 µg/ml of anti-BrdU antibody in PBS-FCS solution (1× PBS, 5% fetal calf serum) for 1 h. After three 3 min washes with PBS, the slides were incubated with 1 µg/ml of fluorescein-labeled anti-mouse IgG for 1 h. The slides were then washed three times for 5 min each with PBS before counterstaining with 4[prime],6-diamidino-2-phenylindole (DAPI) (0.03 µg/ml). The slides were analyzed under a Zeiss fluorescence microscope with a CCD camera.

Flow cytometry and DNA isolation

Two different lymphoblastoid cell lines (GM03715 and GM00546) were cultured and treated with and without aphidicolin at a final concentration of 0.4 µM for 24 h. The cells were then pulse-labeled for 90 min with BrdU at a final concentration of 50 µM. Before sorting, the cells were collected and stained with propidium iodide. The aphidicolin-treated and -untreated BrdU-labeled cells were separated into six different cell cycle phases on a FACS cell sorter. They include G1, four S phases (S1, S2, S3 and S4) and G2/M (30). For each phase, 1 × 105 cells were collected. BrdU-containing DNA from the lymphoblastoid cells and an equal amount of BrdU-labeled DNA from hamster cells were mixed and isolated by co-immunoprecipitation as described previously (27,28,30).

PCR analysis of replicated DNA

A total of 21 PCR-amplifiable sequence-tagged site (STS) markers (see Fig. 4 for the list) were analyzed across the FRA3B region including 14 STS markers previously described (20) and primers specific for seven FHIT exons (exons 3-9) (31). The sequences for the FHIT multiplex PCR primer sets were kindly provided by Dr A.F. Gazdar (University of Texas Southwestern Medical Center). The primers for amplifying hamster DNA as a control were designed based on the HXGPRT gene (accession no. X53080) and the TRANA gene (accession no. X96664). The two hamster-specific primer pairs are: HXGPRT forward, 5[prime]-TTCGGGAAAGAAAGCTGTGA-3[prime]; HXGPRT reverse, 5[prime]-TCTCAAGTAGACAGGAGCCTTCC-3[prime]; TRANA1 forward, 5[prime]-AGCAGTAGAAGAAATACAGGTG-3[prime]; and TRANA1 reverse, 5[prime]-ACCTTGTGGTTTGTTGCTC-3[prime]. In addition, internal controls for given replication profiles include assays of several known early and late replicating human sequences. The human HPRT gene (30) is an early replication marker. DXS297 and DXS7857 are late replication reference markers (28).

For non-polymorphic STS markers, PCR reactions were performed in a volume of 12.5 µl containing 100 µM of each dNTP, 1.5 mM MgCl2, 6.25 pmol of each primer pair, 0.5 U of AmpliGold Taq polymerase (Perkin Elmer-Cetus, Branchburg, NJ) and BrdU-labeled DNA corresponding to 1000 flow-sorted cells in 1× reaction buffer II supplied by the manufacturer. For polymorphic STS markers and FHIT primers, 5 µCi of [[alpha]-32P]dCTP (Amersham) was put into each PCR reaction for direct labeling. All reactions were performed on a Themocycler 480 (Perkin-Elmer Cetus) with 10 min initial denaturation at 95°C to activate the Taq polymerase. Amplification was for 25 cycles which included 94°C for 20 s, 58-60°C for 30 s and 72°C for 30 s. Except for FHIT primer sets, PCR reactions routinely included a pair of hamster primers as a control.

Replication timing analysis

PCR products from non-polymorphic STS markers were run on 1.5% agarose gels, transferred to nylon membranes and then probed with 32P-labeled PCR products. The membrane was reprobed with [alpha]-32P-labeled hamster-specific PCR products. The multiplex PCR products from the FHIT exons were run on 5% non-denaturing polyacrylamide gels. PCR products from polymorphic STS markers were analyzed by 6% denaturing polyacrylamide gels with 8 M urea. The films were exposed for 1-48 h before development. All of the experiments analyzing the cell cycle purified fractions were repeated at least twice.

ACKNOWLEDGEMENTS

The authors thank Dr A.F. Gazdar at the University of Texas Southwestern Medical Center for providing multiplex PCR primer sequences of the FHIT gene and Dr J.Q. Qian at the Mayo Clinic for his technical support for chromosome analysis. This work was supported by NIH grant CA48031 (D.I.S.), DOD grant DAMD-98-1-8522 (D.I.S.) and by the Mayo Foundation.

ABBREVIATIONS

BrdU, 5-bromo-2[prime]-deoxyuridine; FHIT, fragile histidine triad; hRCC, hereditary renal cell carcinoma; LR, late replication.

REFERENCES

1. Smeets, D.F.C.M., Scheres, J.M.J.C. and Hustinx, T.W.J. (1986) The most common fragile site in man is 3p14. Hum. Genet., 72, 215-220. MEDLINE Abstract

2. Brauch, H., Johnson, B., Hovis, J., Yano, T., Gazdar, A., Pettengill, O.S., Graziano, S., Sorenson, G.D., Poiesz, B.J., Minna, J. et al). (1987) Molecular analysis of the short arm of chromosome 3 in small-cell and non-small-cell carcinoma of the lung. N. Engl. J. Med., 317, 1109-1113. MEDLINE Abstract

3. Zbar, B., Brauch, H., Talmadge, C. and Linehan, M. (1987) Loss of alleles of loci on the short arm of chromosome 3 in renal cell carcinoma. Nature, 327, 721-724. MEDLINE Abstract

4. Naylor, S.L., Johnson, B.E., Minna, J.D. and Sakaguchi, A.Y. (1987) Loss of heterozygosity of chromosome 3p markers in small-cell lung cancer. Nature, 329, 451-454. MEDLINE Abstract

5. Shridhar, R., Shridhar, V., Wang, X., Paradee, W., Dugan, M., Sarkar, F., Wilke, C., Glover, T.W., Vaitkevicius, V.K. and Smith, D.I. (1996) Frequent breakpoints in the 3p14.2 fragile site, FRA3B, in pancreatic tumors. Cancer Res., 56, 4347-4350. MEDLINE Abstract

6. Shridhar, V., Wang, L., Rosati, R., Paradee, W., Shridhar, R., Mullins, C., Sakr, W., Grignon, D., Miller, O.J., Sun, Q.C., Petros, J. and Smith, D.I. (1997) Frequent breakpoints in the region surrounding FRA3B in sporadic renal cell carcinomas. Oncogene, 14, 1269-1277. MEDLINE Abstract

7. Ohta, M., Inoue, H., Cotticelli, M.G., Kastury, K., Baffa, R., Palazzo, J., Siprashvili, Z., Mori, M., McCue, P., Druck, T. et al). (1996) The FHIT gene, spanning the chromosome 3p14.2 fragile site and renal carcinoma-associated t(3;8) breakpoint, is abnormal in digestive tract cancers. Cell, 84, 587-597. MEDLINE Abstract

8. Sozzi, G., Veronese, M.L., Negrini, M., Baffa, R., Cotticelli, M.G., Inoue, H., Tornielli, S., Pilotti, S., De Gregorio, L., Pastorino, U., Pierotti, M.A., Ohta, M., Huebner, K. and Croce, C.M. (1996) The FHIT gene 3p14.2 is abnormal in lung cancer. Cell, 85, 17-26. MEDLINE Abstract

9. Virgilio, L., Shuster, M., Gollin, S.M., Veronese, M.L., Ohta, M., Huebner, K. and Croce, C.M. (1996) FHIT gene alterations in head and neck squamous cell carcinomas. Proc. Natl Acad. Sci. USA, 93, 9770-9775.

10. Barnes, L.D., Garrison, P.N., Siprashvili, Z., Guranowski, A., Robinson, A.K., Ingram, S.W., Croce, C.M., Ohta, M. and Huebner, K. (1996) Fhit, a putative tumor suppressor in humans, is a dinucleoside 5[prime], 5[prime][prime][prime]-P1, P3-triphosphate hydrolase. Biochemistry, 35, 11529-11535. MEDLINE Abstract

11. Paradee, W., Wilke, C.M., Wang, L., Shridhar, R., Mullins, C.M,, Hoge, A., Glover, T.W. and Smith, D.I. (1996) A 350-kb cosmid contig in 3p14.2 that crosses the t(3;8) hereditary renal cell carcinoma translocation breakpoint and 17 aphidicolin-induced FRA3B breakpoints. Genomics, 35, 87-93. MEDLINE Abstract

12. Boldog, F., Gemmill, R.M., West, J., Robinson, M., Robinson, L., Li, E., Roche, J., Todd, S., Waggoner, B., Lundstrom, R., Jacobson, J., Mullokandov, M.R., Klinger, H. and Drabkin, H.A. (1997) Chromosome 3p14 homozygous deletions and sequence analysis of FRA3B. Hum. Mol. Genet., 6, 193-203. MEDLINE Abstract

13. Laird, C.D., Jaffe, E., Karpen, G., Lamb, M. and Nelson, R. (1987) Fragile sites in human chromosomes as regions of late replicating DNA. Trends Genet., 3, 274-281.

14. Le Beau, M.M., Rassool, F.V., Neilly, M.E., Espinosa, R. III, Glover, T.W., Smith, D.I. and McKeithan, T.W. (1998) Replication of a common fragile site, FRA3B, occurs late in S phase and is delayed further upon induction: implications for the mechanism of fragile site induction. Hum. Mol. Genet., 7, 755-761. MEDLINE Abstract

15. Glover, T.W., Berger, C. Coyle, J. and Echo, B. (1984) DNA polymerase alpha inhibition by aphidicolin induces gaps and breaks at common fragile sites in human chromosomes. Hum. Genet., 67, 136-142. MEDLINE Abstract

16. Kitsberg, D., Selig, S., Brandeis, M., Simon, I., Keshet, I., Driscoll, D.J., Nicholls, R.D. and Cedar, H. (1993) Allele-specific replication timing of imprinted gene regions. Nature, 364, 459-463. MEDLINE Abstract

17. Lin, M.S., Zhang, A. and Fujimoto, A. (1995) Asynchronous DNA replication between 15q11.2q12 homologs: cytogenetic evidence for maternal imprinting and delayed replication. Hum. Genet., 96, 572-576. MEDLINE Abstract

18. Knoll, J.H., Cheng, S.D. and Lalande, M. (1994) Allele specificity of DNA replication timing in the Angelman/Prader-Willi syndrome imprinted chromosomal region. Nature Genet., 6, 41-46. MEDLINE Abstract

19. Reik, W., Brown, K.W., Schneid, H., Le Bouc, Y., Bickmore, W. and Maher, E.R. (1995) Imprinting mutations in the Beckwith-Wiedemann syndrome suggested by altered imprinting pattern in the IGF2-H19 domain. Hum. Mol. Genet., 4, 2379-2385. MEDLINE Abstract

20. Wang, L., Darling, J., Zhang, J.S., Qian, C.P., Hartmann, L., Conover, C., Jenkins, R. and Smith, D.I. (1998) Frequent homozygous deletions in the FRA3B region in tumor cell lines still leave the FHIT exons intact. Oncogene, 16, 635-642. MEDLINE Abstract

21. Panagopoulos, I., Thelin, S., Mertens, F., Mitelman, F. and Aman, P. (1997) Variable FHIT transcripts in non-neoplastic tissues. Genes Chromosomes Cancer, 19, 215-219. MEDLINE Abstract

22. Luan, X., Shi, G., Zohouri, M., Paradee, W., Smith, D.I., Decker, H.J. and Cannizzaro, L.A. (1997) The FHIT gene is alternatively spliced in normal kidney and renal cell carcinoma. Oncogene, 15, 79-86. MEDLINE Abstract

23. Davies, K. (1992) Imprinting and splicing join together. Nature, 360, 492.

24. Dittrich, B., Buiting, K., Korn, B., Rickard, S., Buxton, J., Saitoh, S., Nicholls, R.D., Poustka, A., Winterpacht, A., Zabel, B. and Horsthemke, B. (1996) Imprint switching on human chromosome 15 may involve alternative transcripts of the SNRPN gene. Nature Genet., 14, 163-170. MEDLINE Abstract

25. Eichler, E.E., Richards, S., Gibbs, R.A. and Nelson, D.L. (1993) Fine structure of the human FMR1 gene. Hum. Mol. Genet., 2, 1147-1153. MEDLINE Abstract

26. Gecz, J., Bielby, S., Sutherland, G.R. and Mulley, J.C. (1997) Gene structure and subcellular localization of FMR2, a member of a new family of putative transcription activators. Genomics, 44, 201-213. MEDLINE Abstract

27. Hansen, R.S., Canfield, T.K., Lamb, M.M., Gartler, S.M. and Laird, C.D. (1993) Association of fragile X syndrome with delayed replication of the FMR1 gene. Cell, 73, 1403-1409. MEDLINE Abstract

28. Hansen, R.S., Canfield, T.K., Fjeld, A.D., Mumm, S., Laird, C.D. and Gartler SM. (1997) A variable domain of delayed replication in FRAXA fragile X chromosomes: X inactivation-like spread of late replication. Proc. Natl Acad. Sci. USA, 94, 4587-4592. MEDLINE Abstract

29. Latos-Bielenska, A., Hameister, H. and Vogel, W. (1987) Detection of BrdUrd incorporation in mammalian chromosomes by a BrdUrd antibody. III. Demonstration of replication patterns in highly resolved chromosomes. Hum. Genet., 76, 293-295. MEDLINE Abstract

30. Hansen, R.S., Canfield, T.K. and Gartler, S.M. (1995) Reverse replication timing for the XIST gene in human fibroblasts. Hum. Mol. Genet., 4, 813-820. MEDLINE Abstract

31. Ahmadian, M., Wistuba, I.I., Fong, K.M., Behrens, C., Kodagoda, D.R., Saboorian, M.H., Shay, J., Tomlinson, G.E., Blum, J., Minna, J.D. and Gazdar, A.F. (1997) Analysis of the FHIT gene and FRA3B region in sporadic breast cancer, preneoplastic lesions, and familial breast cancer probands. Cancer Res., 57, 3664-3668. MEDLINE Abstract

*To whom correspondence should be addressed. Tel: +1 507 266 0311; Fax: +1 507 266 5193; Email: smith.david@mayo.edu

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