Neurofibromatosis type 1 (NF1) is characterized by café-au-lait spots, skinfold freckling, and cutaneous neurofibromas. No obvious relationships between small mutations (<20 bp) of the NF1 gene and a specific phenotype have previously been demonstrated, which suggests that interaction with either unlinked modifying genes and/or the normal NF1 allele may be involved in the development of the particular clinical features associated with NF1. We identified 21 unrelated probands with NF1 (14 familial and 7 sporadic cases) who were all found to have the same c.2970-2972 delAAT (p.990delM) mutation but no cutaneous neurofibromas or clinically obvious plexiform neurofibromas. Molecular analysis identified the same 3-bp inframe deletion (c.2970-2972 delAAT) in exon 17 of the NF1 gene in all affected subjects. The ΔAAT mutation is predicted to result in the loss of one of two adjacent methionines (codon 991 or 992) (ΔMet991), in conjunction with silent ACA→ACG change of codon 990. These two methionine residues are located in a highly conserved region of neurofibromin and are expected, therefore, to have a functional role in the protein. Our data represent results from the first study to correlate a specific small mutation of the NF1 gene to the expression of a particular clinical phenotype. The biological mechanism that relates this specific mutation to the suppression of cutaneous neurofibroma development is unknown. Neurofibromatosis type 1 (NF1) is characterized by café-au-lait spots, skinfold freckling, and cutaneous neurofibromas. No obvious relationships between small mutations (<20 bp) of the NF1 gene and a specific phenotype have previously been demonstrated, which suggests that interaction with either unlinked modifying genes and/or the normal NF1 allele may be involved in the development of the particular clinical features associated with NF1. We identified 21 unrelated probands with NF1 (14 familial and 7 sporadic cases) who were all found to have the same c.2970-2972 delAAT (p.990delM) mutation but no cutaneous neurofibromas or clinically obvious plexiform neurofibromas. Molecular analysis identified the same 3-bp inframe deletion (c.2970-2972 delAAT) in exon 17 of the NF1 gene in all affected subjects. The ΔAAT mutation is predicted to result in the loss of one of two adjacent methionines (codon 991 or 992) (ΔMet991), in conjunction with silent ACA→ACG change of codon 990. These two methionine residues are located in a highly conserved region of neurofibromin and are expected, therefore, to have a functional role in the protein. Our data represent results from the first study to correlate a specific small mutation of the NF1 gene to the expression of a particular clinical phenotype. The biological mechanism that relates this specific mutation to the suppression of cutaneous neurofibroma development is unknown. Neurofibromatosis type 1 (NF1 [MIM 162200]) is a complex disorder that affects many cell types and involves multiple body systems. Inheritance is autosomal dominant, with a prevalence of 1 in 4,0001Huson SM Harper PS Compston DAS Von Recklinghausen neurofibromatosis: clinical and population study in south-east Wales.Brain. 1988; 111: 1355-1381Crossref PubMed Scopus (530) Google Scholar and with penetrance of the mutant gene essentially complete by age 5 years. This inherited disorder is due to germline mutations of the NF1 gene, a 17q11.2-located gene that spans 280 kb, is composed of 61 exons, and encodes a 12-kb mRNA transcript.2Upadhyaya M Cooper DN Neurofibromatosis type 1: from genotype to phenotype. BIOS Publishers, Oxford1998Google Scholar, 3Viskochil DH Gene structure and function.in: Upadhyaya M Cooper DN Neurofibromatosis type 1: from genotype to phenotype. BIOS Publishers, Oxford1998: 39-56Google Scholar Neurofibromin, the NF1 gene product, is a ubiquitously expressed protein that is usually present at low levels in essentially all tissues. This large protein (2,818 aa) exhibits structural and sequence similarity to the mammalian guanine triphosphatase–activating protein (GAP)–related protein family, whose members are evolutionarily conserved. The most highly conserved region of the protein is the neurofibromin GAP-related domain (GRD), which is encoded by exons 20–27a of the NF1 gene and functions by down-regulating Ras activity.4Martin G Viskochil D Bollag G McCabe PC Crosier WJ Haubruck H Conroy L Clark R O’Connell P Cawthon RM et al.The GAP-related domain of the neurofibromatosis type 1 gene product interacts with ras p21.Cell. 1990; 63: 843-849Abstract Full Text PDF PubMed Scopus (704) Google Scholar, 5Cichowski K Jacks T NF1 tumor suppressor gene function: narrowing the GAP.Cell. 2001; 104: 593-604Abstract Full Text Full Text PDF PubMed Scopus (460) Google Scholar Two other domains of neurofibromin have been described—a cysteine/serine–rich domain (CSRD) and a Sec14p domain.6Fahsold R Hoffmeyer S Mischung C Gille C Ehlers C Kucukceylan N Abdel-Nour M Gewies A Peters H Kaufmann D et al.Minor lesion mutational spectrum of the entire NF1 gene does not explain its high mutability but points to a functional domain upstream of the GAP-related domain.Am J Hum Genet. 2000; 66: 790-818Abstract Full Text Full Text PDF PubMed Scopus (234) Google Scholar, 7D’Angelo I Welti S Bonneau Scheffzek K A novel bipartite phospholipids-binding module in neurofibromatosis type 1 protein.EMBO Rep. 2006; 7: 174-179Crossref PubMed Scopus (83) Google Scholar The major clinical features of NF1 are multiple café-au-lait (CAL) spots, skinfold freckling, benign neurofibromas, and Lisch nodules. The development of these features in patients with NF1 is age dependent, with six or more CAL spots usually present by the 2nd year of life, whereas cutaneous neurofibromas develop only later in childhood or in adulthood. Cutaneous neurofibromas are present in almost all patients with NF1 by age 20 years.1Huson SM Harper PS Compston DAS Von Recklinghausen neurofibromatosis: clinical and population study in south-east Wales.Brain. 1988; 111: 1355-1381Crossref PubMed Scopus (530) Google Scholar, 8Friedman JM Birch PH Type 1 neurofibromatosis: a descriptive analysis of the disorder in 1728 patients.Am J Med Genet. 1997; 70: 138-143Crossref PubMed Scopus (313) Google Scholar They lie within the dermis or epidermis and are found to move with the skin when it is manipulated on examination. Neurofibromas may also develop at any point along the course of the peripheral nerves that arise from the spinal root outward. The neurofibromas that develop on the more peripheral nerves are usually palpable as discrete nodular subcutaneous swellings and are usually painful on palpation. These deeper lesions are usually referred to as “subcutaneous” or “nodular” neurofibromas, and they were reported to be present in 27% (81/298) of patients in one study.9Khosrotehrani K Bastji-Garin S Riccardi VM Birch P Friedman JM Wolkenstein P Subcutaneous neurofibromas are associated with mortality in neurofibromatosis type one: a cohort study of 703 patients.Am J Med Genet A. 2005; 132: 49-53Crossref Scopus (55) Google Scholar Only a small proportion of these neurofibromas cause symptoms sufficient to require their removal; only ∼5% (4/86) of adults with NF1 in a Welsh population study had had one removed because of localized paresthesia, whereas symptomatic spinal neurofibromas were removed in only 2% (2/86) of the affected individuals in the same study.1Huson SM Harper PS Compston DAS Von Recklinghausen neurofibromatosis: clinical and population study in south-east Wales.Brain. 1988; 111: 1355-1381Crossref PubMed Scopus (530) Google Scholar, 10Huson SM (1989) Clinical and genetic studies of von Recklinghausen neurofibromatosis. MD thesis, University of Edinburgh, EdinburghGoogle Scholar The other main type of neurofibroma are those referred to as “plexiform” neurofibromas, which can be divided into two types—diffuse and nodular.11Korf B Plexiform neurofibromas.Am J Med Genet. 1999; 89: 31-37Crossref PubMed Scopus (276) Google Scholar Diffuse neurofibromas are the most common form; they are found on routine examination of the skin and usually present as diffuse subcutaneous swellings with ill-defined margins. They were present in 30% of patients in the Welsh series.1Huson SM Harper PS Compston DAS Von Recklinghausen neurofibromatosis: clinical and population study in south-east Wales.Brain. 1988; 111: 1355-1381Crossref PubMed Scopus (530) Google Scholar The overlying dermis and epidermis are often affected by a combination of hypertrophy, pigmentation, and hypertrichosis. Most of these lesions are located on the trunk and are not associated with major problems. Those located on the face, however, may be associated with major disfigurement, and large lesions located elsewhere may be associated with significant bony and/or skin overgrowth. The nodular plexiform neurofibromas may not be obvious on routine examination because they are located more deeply within the body, involving either the major nerve plexuses or the spinal roots. The advent of computed tomography and magnetic resonance imaging scanning has given us the opportunity to study the frequency of these internal tumors in populations with NF1. These studies have shown a high frequency of asymptomatic spinal and internal plexiform lesions.12Thakkar SD Feigen U Mautner VF Spinal tumours in neurofibromatosis type one: an MRI study of frequency, multiplicity and variety.Neuroradiology. 1999; 41: 625-629Crossref PubMed Scopus (117) Google Scholar, 13Schorry EK Crawford AH Egelhoff JC Lovell AM Saal HM Thoracic tumours in children with neurofibromatosis-1.Am J Med Genet. 1997; 74: 533-537Crossref PubMed Scopus (29) Google Scholar, 14Tonsgard JH Kwak SM Short PM Dachman AH CT imaging in adults with neurofibromatosis-1: frequent asymptomatic plexiform lesions.Neurology. 1998; 50: 1755-1760Crossref PubMed Scopus (125) Google Scholar Numerous other disease complications affecting various body systems can occur, and their development usually cannot be predicted, even within families. These include learning problems, various malignant tumors, scoliosis, long-bone pseudarthrosis, and cardiovascular disease in the form of arterial stenosis and pulmonary valve stenosis.1Huson SM Harper PS Compston DAS Von Recklinghausen neurofibromatosis: clinical and population study in south-east Wales.Brain. 1988; 111: 1355-1381Crossref PubMed Scopus (530) Google Scholar, 15Friedman JM Gutmann DH MacCollin M Riccardi VM Neurofibromatosis: phenotype, natural history, and pathogenesis. 3rd ed. The Johns Hopkins University Press, Baltimore and London1999Google Scholar, 16Rasmussen SJ Friedman JM NF1 gene and neurofibromatosis 1.Am J Epidemiol. 2000; 151: 33-40Crossref PubMed Scopus (277) Google Scholar There are a number of minor NF1 features that are not usually associated with significant morbidity but that are encountered at a much greater frequency in patients with NF1 than in the general population. These features include short stature, macrocephaly (both true and relative), and pectus excavatum or carinatum.1Huson SM Harper PS Compston DAS Von Recklinghausen neurofibromatosis: clinical and population study in south-east Wales.Brain. 1988; 111: 1355-1381Crossref PubMed Scopus (530) Google Scholar, 17Cnossen MH Moons KGM Garssen MPJ Pasmans NMT de Goede-Bolder A Niermeijer MF Grobbee DE Minor disease features in neurofibromatosis type 1 (NF1) and their possible value in the diagnosis of NF1 in children ≤6 years and clinically suspected of having NF1.J Med Genet. 1998; 35: 624-627Crossref PubMed Google Scholar Rare families have been reported in which the majority of affected individuals exhibit very similar NF1 features in two or more generations. The best-defined of these families are those with familial CAL spots alone, with or without skinfold freckling.18Charrow J Listernick R Ward K Autosomal dominant multiple café-au-lait spots and neurofibromatosis 1: evidence of non-linkage.Am J Med Genet. 1993; 45: 606-608Crossref PubMed Scopus (44) Google Scholar, 19Brunner HG Hulsebos T Steijlen PM der Kinderen DJ vd Steen A Hamel BC Exclusion of the neurofibromatosis 1 locus in a family with inherited café-au-lait spots.Am J Med Genet. 1993; 46: 472-474Crossref PubMed Scopus (27) Google Scholar, 20Abeliovich D Gelman-Kohan Z Silverstein S Lerer I Chemke J Merin S Zlotogora J Familial café-au-lait spots: a variant of neurofibromatosis type 1.J Med Genet. 1995; 32: 985-986Crossref PubMed Scopus (43) Google Scholar Additional examples include families with Watson syndrome—characterized by pulmonary stenosis, learning problems, and paucity of cutaneous neurofibromas21Allanson JE Upadhyaya M Watson GH Partingon M Mackenzie A Lahey D Macleod H Sarfarazi M Broadhead W Harper PS Watson syndrome: is it a subtype of type 1 neurofibromatosis?.J Med Genet. 1991; 28: 752-756Crossref PubMed Scopus (76) Google Scholar, 22Upadhyaya M Shen M Cherryson A Farnham J Maynard J Huson SM Harper PS Analysis of mutations in neurofibromatosis type 1.Hum Mol Genet. 1992; 1: 735-740Crossref PubMed Scopus (76) Google Scholar, 23Tassabehji M Strachan T Sharland M Colley A Donnai D Harris R Thakker N Tandem duplication within a neurofibromatosis type 1 (NF1) gene exon in a family with features of Watson syndrome and Noonan syndrome.Am J Hum Genet. 1993; 53: 90-95PubMed Google Scholar—and families with multiple spinal neurofibromas.24Pulst SM Riccardi VM Fain P Korenberg JR Familial spinal neurofibromatosis: clinical and DNA linkage analysis.Neurology. 1991; 41: 1923-1927Crossref PubMed Google Scholar, 25Poyhonen M Leisti E-L Kytola S Leisti J Hereditary spinal neurofibromatosis: a rare form of NF1?.J Med Genet. 1997; 34: 184-187Crossref PubMed Scopus (51) Google Scholar Of these various familial subtypes, only Watson syndrome appears to be genetically homogeneous but exhibits allelic heterogeneity. Families have been reported with both CAL spots and spinal neurofibromas that are linked or unlinked to the NF1 gene.18Charrow J Listernick R Ward K Autosomal dominant multiple café-au-lait spots and neurofibromatosis 1: evidence of non-linkage.Am J Med Genet. 1993; 45: 606-608Crossref PubMed Scopus (44) Google Scholar, 19Brunner HG Hulsebos T Steijlen PM der Kinderen DJ vd Steen A Hamel BC Exclusion of the neurofibromatosis 1 locus in a family with inherited café-au-lait spots.Am J Med Genet. 1993; 46: 472-474Crossref PubMed Scopus (27) Google Scholar, 20Abeliovich D Gelman-Kohan Z Silverstein S Lerer I Chemke J Merin S Zlotogora J Familial café-au-lait spots: a variant of neurofibromatosis type 1.J Med Genet. 1995; 32: 985-986Crossref PubMed Scopus (43) Google Scholar, 24Pulst SM Riccardi VM Fain P Korenberg JR Familial spinal neurofibromatosis: clinical and DNA linkage analysis.Neurology. 1991; 41: 1923-1927Crossref PubMed Google Scholar, 25Poyhonen M Leisti E-L Kytola S Leisti J Hereditary spinal neurofibromatosis: a rare form of NF1?.J Med Genet. 1997; 34: 184-187Crossref PubMed Scopus (51) Google Scholar In our clinical experience, such families represent <5% of an NF1 clinical population. The majority of families exhibit wide intrafamilial variation in both the number of major features and the occurrence of complications. Two studies quantitatively analyzed the familial variation in NF126Easton DF Ponder MA Huson SM Ponder BA An analysis of variation in expression of neurofibromatosis type 1 (NF1): evidence for modifying genes.Am J Hum Genet. 1993; 53: 305-313PubMed Google Scholar, 27Szudek J Joe H Friedman JM Analysis of intrafamilial phenotypic variation in neurofibromatosis type 1 (NF1).Genet Epidemiol. 2002; 23: 150-164Crossref PubMed Scopus (65) Google Scholar and found evidence for the involvement of modifying genes, and perhaps also the normal NF1 allele, in the development of particular disease features. Subsequent to the cloning of the NF1 gene in 1990, numerous studies have looked at genotype-phenotype correlation in NF1.28Easton DF Genotype-phenotype relationships in neurofibromatosis type 1.in: Upadhyaya M Cooper DN Neurofibromatosis type 1: from genotype to phenotype. BIOS Publishers, Oxford1998: 167-171Google Scholar, 29Castle B Baser ME Huson SM Cooper DN Upadhyaya M Evaluation of genotype-phenotype correlations in neurofibromatosis type 1.J Med Genet. 2003; 40: e109Crossref PubMed Scopus (75) Google Scholar However, with the exception of those patients with the NF1 microdeletion syndrome who develop a large number of neurofibromas for their age, exhibit dysmorphic features, show a probable increased predisposition to the development of malignant peripheral nerve–sheath tumors (MPNST), and exhibit cardiac abnormalities and overgrowth,30Kayes LM Burke W Riccardi VM Bennett R Ehrlich P Rubenstein A Stephens K Deletions spanning the neurofibromatosis 1 gene: identification and phenotype of five patients.Am J Hum Genet. 1994; 54: 424-436PubMed Google Scholar, 31Wu BL Austin MA Schneider GH Boles RG Korf BR Deletion of the entire NF1 gene detected by FISH: four deletion patients associated with severe manifestations.Am J Med Genet. 1995; 59: 528-535Crossref PubMed Scopus (79) Google Scholar, 32Cnossen MH van der Est MN Breuning MH Deletions spanning the neurofibromatosis type 1 gene: correlations in neurofibromatosis type 1?.Hum Mut. 1997; 9: 458-464Crossref PubMed Scopus (91) Google Scholar, 33Leppig KA Kaplan P Viskochil D Weaver M Ortenberg J Stephens K Familial neurofibromatosis 1 microdeletions: cosegregation with distinct facial phenotype and early onset of cutaneous neurofibromata.Am J Med Genet. 1997; 73: 197-204Crossref PubMed Scopus (86) Google Scholar, 34Tonsgard JH Yelavarthi KK Cushner S Short MP Lindgren V Do NF1 gene deletions result in a characteristic phenotype?.Am J Med Genet. 1997; 73: 80-86Crossref PubMed Scopus (99) Google Scholar, 35Lopez-Correa C Dorscher M Brems H Lazaro C Clementi M Upadhyaya M Dooijes D Moog U Kehrer-Sawatzki H Rutkowski JL et al.Recombination hotspot in NF1 microdeletion patients.Hum Mol Genet. 2001; 10: 1387-1392Crossref PubMed Scopus (141) Google Scholar, 36Upadhyaya M Osborn M Maynard J Kim MR Tamanoi F Cooper DN Mutational and functional analysis of NF1.Hum Genet. 1997; 99: 88-92Crossref PubMed Scopus (84) Google Scholar, 37Upadhyaya M Ruggieri M Maynard J Osborn M Hartog C Mudd S Penttinen M Cordeiro I Ponder M Ponder BA et al.Gross deletions of the neurofibromatosis type 1 (NF1) gene are predominantly of maternal origin and commonly associated with a learning disability, dysmorphic features and developmental delay.Hum Genet. 1998; 102: 591-597Crossref PubMed Scopus (133) Google Scholar, 38De Raedt T Brems H Wolkenstein P Vidaud D Pilotti S Perrone F Mautner V Frahm S Sciot R Legius E Elevated risk for MPNST in NF1 microdeletion patients.Am J Hum Genet. 2003; 72: 1288-1292Abstract Full Text Full Text PDF PubMed Scopus (193) Google Scholar, 39Upadhyaya M Spurlock G Majounie E Griffiths S Forrester N Baser M Huson SM Evans G Ferner R The heterogeneous nature of germline NF1 gene mutations in NF1 patients with malignant peripheral nerve sheath tumours (MPNSTs).Hum Mut. 2006; 27: 716Crossref PubMed Scopus (31) Google Scholar, 40Venturin M Guarnieri P Natacci F Stabile M Tenconi R Clementin M Hernandez C Thompson P Upadhyaya M Larizza L et al.Mental retardation and cardiovascular malformations in NF1 microdeleted patients point to candidate genes in 17q11.2.J Med Genet. 2004; 41: 35-41Crossref PubMed Scopus (91) Google Scholar, 41Spiegel M Oexle K Horn D Windt E Buske A Albrecht B Prott E Seemanova E Seidel J Rosenbaum T et al.Childhood overgrowth in patients with common NF1 microdeletions.Eur J Hum Genet. 2005; 13: 883-888Crossref PubMed Scopus (36) Google Scholar no other clinically significant findings have been reported. The finding of the same ΔAAT mutation in exon 17 of the NF1 gene in three families with no neurofibromas prompted us to review our own mutation database for patients with the same mutation and to contact colleagues in other laboratories worldwide. We here report a cohort of 21 unrelated probands, along with an additional 26 affected relatives, all of whom have the same inframe AAT deletion but no cutaneous neurofibromas. Samples from the probands of the three original large multigeneration families (fig. 1A–1C) were referred to the DNA service laboratory in Cardiff because of their unusual phenotypes. Having identified the same mutation in these families, we examined the clinical phenotypes exhibited by four other patients whom we had previously identified with this specific genotype, and we also contacted relevant laboratories worldwide that had extensive experience of NF1 genotyping, to ascertain other patients with this genotype. This study was approved by the appropriate institutional review board. A clinical questionnaire was completed for each patient. The clinical data on pulmonic stenosis were based on the clinician declaring lack of or presence of pulmonic stenosis. The significance of clinical findings was calculated using the Z test for proportions. The one-tailed probabilities associated with the resulting Z values were used to estimate the significance of the difference. Sporadic cases of NF1 who were younger than 6 years were excluded from analysis because, at their age, they would not be expected to have developed cutaneous neurofibromas. NF1 mutation screening in these families involved a variety of complementary mutation-detection techniques, including SSCP, heteroduplex analysis, denaturing high-performance liquid chromatography, direct sequencing, RT-PCR, and multiplex ligation-dependent probe amplification (MLPA).42Ars E Kruyer H Morell M Pros E Serra E Ravella A Estivill X Lazaro C Recurrent mutations in the NF1 gene are common among neurofibromatosis type 1 patients.J Med Genet. 2003; 40: e82Crossref PubMed Scopus (129) Google Scholar, 43Messiaen L Callens T Mortier G Beysen D Vandenbroucke I Van Roy N Speleman F Paepe AD Exhaustive mutation analysis of the NF1 gene allows identification of 955 of mutations and reveals a high frequency of unusual splicing defects.Hum Mut. 2000; 15: 541-555Crossref PubMed Scopus (382) Google Scholar, 44Mattocks C Baralle D Tarpey P ffrench-Constant C Bobrow M Whittaker J Automated comparative sequence analysis identifies mutations in 89% of NF1 patients and confirms a mutation cluster in exons 11-17 distinct from the GAP-related domain.J Med Genet. 2004; 41: e48Crossref PubMed Scopus (57) Google Scholar, 45Upadhyaya M Han S Consoli C Majounie E Horan M Thomas NS Potts C Griffiths S Ruggieri M von Deimling A et al.Characterisation of the somatic mutational spectrum of the neurofibromatosis type 1 (NF1) gene in neurofibromatosis patients with benign and malignant tumours.Hum Mut. 2004; 23: 134-146Crossref PubMed Scopus (80) Google Scholar, 46Griffiths S Thompson P Frayling I Upadhyaya M Molecular diagnosis of neurofibromatosis type 1: 2 years experience.Fam Cancer. 2006; (electronically published August 31, 2006; accessed November 21, 2006)(http://www.springerlink.com/content/vq717jw5u0x25344/?p=1cc7aaa399944ac9a599b5cba9839c46&pi=8)Google Scholar, 47Wimmer K Yao S Claes K Kehrer-Sawatzki H Tinschert S De Raedt T Legius E Callens T Beiglbock H Maertens O et al.Spectrum of single and multi-exon NF1 copy number changes in a cohort of 1,100 unselected NF1 patients.Genes Chrom Cancer. 2006; 45: 265-276Crossref PubMed Scopus (109) Google Scholar To assess the potential contribution that the local DNA sequence structure might have made to the molecular mechanism that produced this specific NF1 microdeletion, we performed complexity analysis of the immediate flanking DNA sequence.48Gusev VD Nemytikova LA Chuzhanova NA On the complexity measures of genetic sequences.Bioinformatics. 1999; 15: 994-999Crossref PubMed Scopus (121) Google Scholar, 49Chuzhanova NA Anassis EJ Ball E Krawczak M Cooper DN Meta-analysis of indels causing human genetic disease: mechanisms of mutagenesis and the role of local DNA sequence complexity.Hum Mut. 2003; 21: 28-44Crossref PubMed Scopus (91) Google Scholar In essence, complexity analysis involves the partitioning of a given sequence S into r consecutive fragments: S=S(1:i1)S(i1+1:i2)…S(ir−1+1:ir=N), where each S(ik−1+1:ik) is the longest fragment downstream of position ik−1 for which a direct, inverted, or symmetric sequence copy is located upstream of position ik−1+1 and where N is the length of the sequence S. For example, if S=ACACCACTCC, then the partition of S computed with respect to direct repeats is A-C-AC-CAC-T-CC. The first, second, and fifth fragments are of length 1, since nucleotides A, C, and T do not occur upstream of their first-occurring positions. The third fragment (AC) is a copy of substring S(1:2), the fourth fragment (CAC) is a copy of substring S(2:4)=CAC, and the sixth fragment (CC) is a copy of substring S(4:5). The number of fragments in the partition, C(S)=r, is termed “the complexity of S.” Thus, for the sequence in the example given above, the complexity with respect to direct repeats is 6. In a similar way, sequence complexity can be calculated with respect to inverted repeats and symmetric elements. A previous complexity analysis of ∼4,000 microdeletions and ∼2,000 microinsertions derived from the Human Gene Mutation Database (HGMD) demonstrated that, in 80%–90% of such mutations, repetitive elements identified as making the most significant contribution to the change in sequence complexity, consequent to a mutation, were of those sequence repeats that were also directly involved in mediating the mutational change itself. Hence, the most probable mechanism of mutation is that in which the change in complexity is maximized.49Chuzhanova NA Anassis EJ Ball E Krawczak M Cooper DN Meta-analysis of indels causing human genetic disease: mechanisms of mutagenesis and the role of local DNA sequence complexity.Hum Mut. 2003; 21: 28-44Crossref PubMed Scopus (91) Google Scholar, 50Ball EV Stenson PD Krawczak M Cooper DN Chuzhanova NA Micro-deletions and micro-insertions causing human genetic disease: common mechanisms of mutagenesis and the role of local DNA sequence complexity.Hum Mut. 2005; 26: 205-213Crossref PubMed Scopus (121) Google Scholar The study group consisted of 21 unrelated probands and 26 affected relatives. The NF1 clinical features exhibited by these 47 affected individuals are summarized in table 1, and pedigrees for the five families in which at least 2 affected individuals were studied are shown in figure 1. The ages of the 47 individuals (21 males and 26 females) examined ranged from 2 to 65 years: 2 were younger than 5 years, 3 were aged 6–10 years, 25 were aged 11–20 years, 2 were aged 21–30 years, 7 were aged 31–40 years, and 8 were older than 40 years.Table 1Clinical Details for 47 Affected Individuals from 21 Families with NF1 with c.2970-2972 delAATClinical FeatureFamily and IndividualSex/ Age (years)Axillary/Inguinal FrecklingLisch NodulesShort StatureMacrocephalyPectus AbnormalityScoliosisPulmonary StenosisLDaLD = learning difficulty.OtherA: 1F/59YYNNNNNNN 2M/65NNNNNNNNMultiple lipomas 3F/56YNNNNNNNN 4F/35YNNNNNNNMultiple lipomas 5M/36YNNYNNNNN 6M/5YNYNNNNYN 7M/2YYNNNNNNNB: 1M/50YNNYNNNNN 2F/14YNNNYYNYArterial stenosis 3F/15YNNNYYNNNC: 1F/63YNN?NNNNN 2F/41YNNNNNNNN 3F/40YNNNNNNNN 4M/17YNNNNNNNN 5F/16YNNNNNNNN 6F/8YNNNNNNYCerebellar cavernous hemangioma 7F/16YNNNNNNNN 8F/14YNNNNNNNN 9F/13YNNNNNNNN 10F/8YNNNNNNNND: 1F/38NNNNNNNNN 2F/11NNNNNNNNN 3F/11NNNNNNNNN 4M/17NNNNNNNNNE: 1F/36Y?NNNNNNN 2F/14N?YNYNNNN 3M/13Y?YNYNYYN 4M/10N?NNYNYYN 5F/8N?YNYNYNNF: 1M/53N?NNNNNNN 2M/19YYNNNNNYAtrial septal defectG: 1M/51YNN??NNNN 2M/20YNN??NNNNH: 1bSporadic cases of NF1. All other individuals were familial cases.M/15NNNNNNNNNI: 1M/34NNNNNYNNNJ: 1bSporadic cases of NF1. All other individuals were familial cases.F/18YNNYNNNNNK: 1F/8YNNNNNNNNL: 1M/12YNNNNNNNNM: 1bSporadic cases of NF1. All other individuals were familial cases.M/11NNYNNNNYGenitourinary abnormalitiesN: 1F/12N?NNYNYNNO: 1M/17Y?NYNNNYNP: 1bSporadic cases of NF1. All other individuals were familial cases.M/15YNNNNNNNNQ: 1bSporadic cases of NF1. All other individuals were familial cases.M/13NNNNNNNNNR: 1M/36NNNNNNNNNS: 1F/10YNNNNNNNNT: 1bSporadic cases of NF1. All other individuals were familial cases.F/27Y?NNNYNYSpinal neurofibromaU: 1bSporadic cases of NF1. All other individuals were familial cases.F/25YNNNNNNNNNote.—All individuals had no cutaneous, subcutaneous, or clinically obvious plexiform neurofibromas. All had six or more CAL macules, except for patient A3.a LD = learning difficulty.b Sporadic cases of NF1. All other individuals were familial cases. Open table in a new tab Note.— All individuals had no cutaneous, subcutaneous, or clinically obvious plexiform neurofibromas. All had six or more CAL macules, except for patient A3. The individuals in families A–G were all examined at the time of data collection for the study. Family E was recently reported because of its NF1 and Noonan syndrome (NS) phenotypes.51Stevenson DA Viskochil DH Rope AF Carey JC Clinical and molecular aspects of an informative family with neurofibromatosis type 1 and Noonan phenotype.Clin Genet. 2006; 69: 246-253Crossref PubMed Scopus (30) Google Scholar The DNA samples from the remaining probands had been previously genotyped in various NF1 research laboratories, and only a few of these patients were subsequently followed up for this study. The proband in family N was originally referred to our laboratory with a diagnosis of Watson syndrome, and this individual was included in a previous study.29Castle B Baser ME Huson SM Cooper DN Upadhyaya M Evaluation of genotype-phenotype correlations in neurofibromatosis type 1.J Med Genet. 2003; 40: e109Crossref PubMed Scopus (75) Google Scholar All affected individuals analyzed
