Microarray analysis detects novel Pax3 downstream target genes.

Pax3 is a transcription factor that is required for the development of embryonic neural tube, neural crest, and somatic derivatives. Our previous study (Mayanil, C. S. K., George, D., Mania-Farnell, B., Bremer, C. L., McLone, D. G., and Bremer, E. G. (2000) J. Biol. Chem. 275, 23259-23266) reveals that overexpression of Pax3 in a human medulloblastoma cell line, DAOY, resulted in an up-regulation in alpha-2,8-polysialyltransferase (STX) gene expression and an increase in polysialic acid on neural cell adhesion molecule. This finding suggests that STX might be a previously undescribed downstream target of Pax3. Because Pax3 is important in diverse cellular functions during development, we are interested in the identification of additional downstream targets of Pax3. We utilized oligonucleotide arrays and RNA isolated from stable Pax3 transfectants to identify potential target genes. A total of 270 genes were altered in the Pax3 transfectants as compared with the vector control and parental cell line. An independent analysis by cDNA expression array and real-time quantitative polymerase chain reaction of several genes confirmed the changes observed by the oligonucleotide microarray data. Of the genes that displayed significant changes in expression, several contain paired and homeodomain binding motifs of Pax3 in their promoter regions. Using promoter-luciferase reporter transfection assays and electromobility shift assays, we showed at least one previously undescribed downstream target, STX, to be a biological downstream target of Pax3. Thus we report several previously undescribed candidate genes to be potential downstream targets of Pax3.

Spina bifida outcome: a 25-year prospective.

BACKGROUND: Open spina bifida is the most complex congenital abnormality compatible with long-term survival. This report outlines the 20- to 25-year outcome for our original cohort of patients with a myelomeningocele treated in a nonselective, prospective manner. METHODS: Of the initial 118 children, 71 patients were available for our most recent review. Nineteen patients have been lost to follow-up and 28 patients have died. Data were collected on: motor level, shunt status, education/employment, seizure history, mobility, bladder/bowel continence, tethered cord, scoliosis, latex allergy, posterior cervical decompression, tracheostomy and/or gastrostomy tube. RESULTS: Mortality (24%) continues to climb into young adulthood. Eighty-six percent of the cohort have cerebrospinal fluid diversion, with 95% having undergone at least one shunt revision. Thirty-two percent have undergone a tethered cord release, with 97% having an improvement or stabilization in their preoperative symptoms. Forty-nine percent have scoliosis, with 43% eventually requiring a spinal fusion. Sixteen patients (23%) have had at least one seizure. Eighty-five percent are attending or have graduated from high school and/or college. More than 80% of young adults have social bladder continence. Approximately 1/3 of patients are allergic to latex, with 6 patients having experienced a life-threatening reaction. CONCLUSION: At least 75% of children born with a myelomeningocele can be expected to reach their early adult years. Late deterioration is common. One of the greatest challenges in medicine today is establishing a network of care for these adults with spina bifida.

Overexpression of murine Pax3 increases NCAM polysialylation in a human medulloblastoma cell line.

Polysialic acid (PSA) is a developmentally regulated carbohydrate found primarily on neural cell adhesion molecules (NCAM) in embryonic tissues. The majority of NCAM in adult tissues lacks this unique carbohydrate, but polysialylated NCAM (PSA-NCAM) is present in adult brain regions where neural regeneration persists and in some pediatric brain tumors such as medulloblastoma, which show greater propensity for leptomeningeal spread. Pax3, a developmentally regulated paired homeodomain transcription factor, is thought to be involved in the regulation of neural cell adhesion molecules. Overexpression of murine Pax3 into a human medulloblastoma cell line (DAOY) resulted in an increase in NCAM polysialylation and a 2-4-fold increase in alpha2, 8-polysialyltransferase type II mRNA levels. No difference was observed in alpha2,8-polysialyltransferase type IV message. The addition of PSA to NCAM changed the adhesive behavior of these Pax3 transfectants. Transfectants expressing high PSA-NCAM show much less NCAM-dependent aggregation than those with less PSA-NCAM. In addition, Pax3 transfectants having high PSA-NCAM show heterophilic adhesion involving polysialic acid to heparan sulfate proteoglycan and agrin. These observations suggest that a developmentally regulated transcription factor, Pax3, could affect NCAM polysialylation and subsequently cell-cell and cell-substratum interaction.

Myelocystocele-cloacal exstrophy in a pedigree with a mitochondrial 12S rRNA mutation, aminoglycoside-induced deafness, pigmentary disturbances, and spinal anomalies.

A large Filipino-American family with progressive matrilineal hearing loss, premature graying, depigmented patches, and digital anomalies was ascertained through a survey of a spina bifida clinic for neural crest disorders. Deafness followed a matrilineal pattern of inheritance and was associated with the A1555G mutation in the 12S rRNA gene (MTRNR1) in affected individuals as well as unaffected maternal relatives. Several other malformations were found in carriers of the mutation. The proband had a myelocystocele, Arnold-Chiari type I malformation, cloacal exstrophy, and severe early-onset hearing loss. Several family members had premature graying, white forelock, congenital leukoderma with or without telecanthus, somewhat suggestive of a Waardenburg syndrome variant. In addition to the patient with myelocystocele, two individuals had scoliosis and one had segmentation defects of spinal vertebrae. The syndromic characteristics reported here are novel for the mitochondrial A1555G substitution, and may result from dysfunction of mitochondrial genes during early development. However, the mitochondrial A1555G mutation is only rarely associated with neural tube defects as it was not found in a screen of 218 additional individuals with spina bifida, four of whom had congenital hearing loss.

Neural crest anomaly syndromes in children with spina bifida.

A hereditary contribution to the etiology of neural tube defects (NTDs) has been suggested by clinical studies and animal models. To evaluate the hypothesis that common genes are important for both neural tube defects and neural crest anomalies, we examined children with developmental abnormalities of the spinal cord for anomalies of neural crest-derived structures. Neural crest anomalies, particularly auditory and pigmentary disorders, were identified and classified according to inheritance and type of anomaly. Of the 515 children screened, 44 (8.5%) had neural crest anomalies, 20 (3.9%) of which were apparently familial. Another 19 (3.7%) families had neural crest anomalies in two or more close relations, but the NTD subject was unaffected. Sixteen (3.1%) children with NTDs had a recognizable syndrome, including nine (1.7%) with a subtype of the Waardenburg syndromes. The coincidence of familial neural crest anomaly syndromes in subjects with spina bifida implies that defects in genes underlying neural crest development may contribute to the etiology of neural tube defects in a fraction of cases. The rate of anomalies and familial syndromes of neural crest-derived structures must be assessed in an adequate control sample to evaluate whether or not these abnormalities constitute risk factors for NTDs.

The presence of transcription factors in chicken albumin, yolk and blastoderm.

Embryonic development is determined by preset intrinsic programs and extrinsic signals. To explore the possibility that transcription factors are present at the onset of development, preparations of yolk, albumin, and blastoderm from unfertilized and fertilized white Leghorn chicken eggs were screened by a panel of 16 transcription factor antibodies with Western blot techniques. Yolk was positive for 13 transcription factors, whereas blastoderm was positive for 10, and albumin was positive for 5. In yolk, several transcription factors, GATA-2, E2F-1, MyoD, and TFIID, were developmentally regulated. These results indicate that intracellular yolk and extracellular albumin contain transcription factors which presumably influence early chick embryonic development from prefertilization to the late blastoderm stage. Thus, the utility of preset maternal transcription factors within yolk and albumin complement maternally derived mRNA to determine the early development of the zygote.

The biological resolution of malformations of the central nervous system.

It has been known for years that the assembly of the nervous system is under genetic control. During the last 10 years, the genes that direct the formation of the brain and spinal cord have begun to be discovered at an amazing pace. Mutations in the fruit fly and advances in molecular genetics have led the way. Gene mutations that cause many of the malformations of the human brain and spinal cord are now known. This has many physician-scientists hoping that an understanding of cause might lead to cure.