Nasal Nitric Oxide in Primary Immunodeficiency and Primary Ciliary Dyskinesia: Helping to Distinguish Between Clinically Similar Diseases.

PURPOSE: Primary ciliary dyskinesia (PCD) is a rare disorder of the mucociliary clearance leading to recurrent upper and lower respiratory tract infections. PCD is difficult to clinically distinguish from other entities leading to recurrent oto-sino-pulmonary infections, including primary immunodeficiency (PID). Nasal nitric oxide (nNO) is a sensitive and specific diagnostic test for PCD, but it has not been thoroughly examined in PID. Past publications have suggested an overlap in nNO levels among subjects with PCD and PID. We sought to determine if nNO measurements among patients diagnosed with PID would fall significantly above the established PCD diagnostic cutoff value of 77 nL/min. METHODS: Children > 5 years old and adults with definitive PID or PCD diagnoses were recruited from outpatient subspecialty clinics. Participants underwent nNO testing by standardized protocol using a chemiluminescence analyzer and completed a questionnaire concerning their chronic oto-sino-pulmonary symptoms, including key clinical criteria specific to diagnosed PCD (neonatal respiratory distress at term birth, year-round cough or nasal congestion starting before 6 months of age, any organ laterality defect). RESULTS: Participants included 32 patients with PID, 27 patients with PCD, and 19 healthy controls. Median nNO was 228.9.1 nL/min in the PID group, 19.7 nL/min in the PCD group, and 269.4 in the healthy controls (p < 0.0001). Subjects with PCD were significantly more likely to report key clinical criteria specific to PCD, but approximately 25% of PID subjects also reported at least 1 of these key clinical criteria (mainly year-round cough or nasal congestion). CONCLUSIONS: While key clinical criteria associated with PCD often overlap with the symptoms reported in PID, nNO measurement by chemiluminescence technology allows for effective discrimination between PID and PCD.

Primary Ciliary Dyskinesia: Longitudinal Study of Lung Disease by Ultrastructure Defect and Genotype.

RATIONALE: In primary ciliary dyskinesia, factors leading to disease heterogeneity are poorly understood. OBJECTIVES: To describe early lung disease progression in primary ciliary dyskinesia and identify associations between ultrastructural defects and genotypes with clinical phenotype. METHODS: This was a prospective, longitudinal (5 yr), multicenter, observational study. Inclusion criteria were less than 19 years at enrollment and greater than or equal to two annual study visits. Linear mixed effects models including random slope and random intercept were used to evaluate longitudinal associations between the ciliary defect group (or genotype group) and clinical features (percent predicted FEV1 and weight and height z-scores). MEASUREMENTS AND MAIN RESULTS: A total of 137 participants completed 732 visits. The group with absent inner dynein arm, central apparatus defects, and microtubular disorganization (IDA/CA/MTD) (n = 41) were significantly younger at diagnosis and in mixed effects models had significantly lower percent predicted FEV1 and weight and height z-scores than the isolated outer dynein arm defect (n = 55) group. Participants with CCDC39 or CCDC40 mutations (n = 34) had lower percent predicted FEV1 and weight and height z-scores than those with DNAH5 mutations (n = 36). For the entire cohort, percent predicted FEV1 decline was heterogeneous with a mean (SE) decline of 0.57 (0.25) percent predicted/yr. Rate of decline was different from zero only in the IDA/MTD/CA group (mean [SE], -1.11 [0.48] percent predicted/yr; P = 0.02). CONCLUSIONS: Participants with IDA/MTD/CA defects, which included individuals with CCDC39 or CCDC40 mutations, had worse lung function and growth indices compared with those with outer dynein arm defects and DNAH5 mutations, respectively. The only group with a significant lung function decline over time were participants with IDA/MTD/CA defects.

Yeast mRNA Poly(A) tail length control can be reconstituted in vitro in the absence of Pab1p-dependent Poly(A) nuclease activity.

Regulation of poly(A) tail length during mRNA 3'-end formation requires a specific poly(A)-binding protein in addition to the cleavage/polyadenylation machinery. The mechanism that controls polyadenylation in mammals is well understood and involves the nuclear poly(A)-binding protein PABPN1. In contrast, poly(A) tail length regulation is poorly understood in yeast. Previous studies have suggested that the major cytoplasmic poly(A)-binding protein Pab1p acts as a length control factor in conjunction with the Pab1p-dependent poly(A) nuclease PAN, to regulate poly(A) tail length in an mRNA specific manner. In contrast, we recently showed that Nab2p regulates polyadenylation during de novo synthesis, and its nuclear location is more consistent with a role in 3'-end processing than that of cytoplasmic Pab1p. Here, we investigate whether PAN activity is required for de novo poly(A) tail synthesis. Components required for mRNA 3'-end formation were purified from wild-type and pan mutant cells. In both situations, 3'-end formation could be reconstituted whether Nab2p or Pab1p was used as the poly(A) tail length control factor. However, polyadenylation was more efficient and physiologically more relevant in the presence of Nab2p as opposed to Pab1p. Moreover, cell immunofluorescence studies confirmed that PAN subunits are localized in the cytoplasm which suggests that cytoplasmic Pab1p and PAN may act at a later stage in mRNA metabolism. Based on these findings, we propose that Nab2p is necessary and sufficient to regulate poly(A) tail length during de novo synthesis in yeast.

Dual requirement for yeast hnRNP Nab2p in mRNA poly(A) tail length control and nuclear export.

Recent studies of mRNA export factors have provided additional evidence for a mechanistic link between mRNA 3'-end formation and nuclear export. Here, we identify Nab2p as a nuclear poly(A)-binding protein required for both poly(A) tail length control and nuclear export of mRNA. Loss of NAB2 expression leads to hyperadenylation and nuclear accumulation of poly(A)(+) RNA but, in contrast to mRNA export mutants, these defects can be uncoupled in a nab2 mutant strain. Previous studies have implicated the cytoplasmic poly(A) tail-binding protein Pab1p in poly(A) tail length control during polyadenylation. Although cells are viable in the absence of NAB2 expression when PAB1 is overexpressed, Pab1p fails to resolve the nab2Delta hyperadenylation defect even when Pab1p is tagged with a nuclear localization sequence and targeted to the nucleus. These results indicate that Nab2p is essential for poly(A) tail length control in vivo, and we demonstrate that Nab2p activates polyadenylation, while inhibiting hyperadenylation, in the absence of Pab1p in vitro. We propose that Nab2p provides an important link between the termination of mRNA polyadenylation and nuclear export.