Coordination of phytochrome levels in phyB mutants of Arabidopsis as revealed by apoprotein-specific monoclonal antibodies.

Accumulating evidence indicates that individual members of the phytochrome family of photoreceptors have differential but interactive roles in controlling plant responses to light. To investigate possible cross-regulation of these receptors, we have identified monoclonal antibodies that specifically detect each of the five Arabidopsis phytochromes, phyA to phyE (phytochrome A holoprotein; PHYA, phytochrome A apoprotein; PHYA, phytochrome A gene; phyA, mutant allele of phytochrome A gene), on immunoblots and have used them to analyze the effects of phyA and phyB null mutations on the levels of all five family members. In phyB mutants, but not in phyA mutants, a four- to six-fold reduction in the level of phyC is observed in tissues grown either in the dark or in the light. Coordinate expression of phyB and phyC is induced in the phyB mutant background by the presence of a complementing PHYB transgene. However, in transgenic lines that overexpress phyB 15- to 20-fold, phyC is not similarly overexpressed. In these overexpressor lines, the levels of phyA, phyC, and phyD are increased two- to four-fold over normal in light-grown but not dark-grown seedlings. These observations indicate that molecular mechanisms for coordination or cross-regulation of phytochrome levels are active in Arabidopsis and have implications for the interpretation of phytochrome mutants and overexpressor lines.

A deletion in the PHYD gene of the Arabidopsis Wassilewskija ecotype defines a role for phytochrome D in red/far-red light sensing.

The PHYD gene of the Wassilewskija (Ws) ecotype of Arabidopsis contains a 14-bp deletion (the phyD-1 mutation) beginning at amino acid 29 of the reading frame, resulting in translation termination at a nonsense codon 138 nucleotides downstream of the deletion end point. Immunoblot analyses showed that Ws lacks phyD but contains normal levels of phyA, phyB, and phyC. By backcrossing into the Ws and Landsberg erecta genetic backgrounds, we constructed sibling pairs of PHYD+ and phyD-1 lines and of phyB- PHYD+ and phyB- phyD- lines. Hypocotyl lengths after growth under white or red light increased sequentially in strains that were B+D+, B+D-, B-D+, and B-D-. In the Ws genetic background, an increase in petiole length, a reduction in cotyledon area and in anthocyanin accumulation in seedling stems, a diminished effect of an end-of-day pulse of far-red light on hypocotyl elongation, and a decrease in the number of rosette leaves at the onset of flowering were also seen sequentially in these lines. Thus, phyD, which is approximately 80% identical in amino acid sequence to phyB, acts in conjunction with phyB in regulating many shade avoidance responses. The existence of the apparently naturally occurring phyD-1 mutation indicates that phyD is not essential in some natural environments.

The phytochrome apoprotein family in Arabidopsis is encoded by five genes: the sequences and expression of PHYD and PHYE.

Two novel Arabidopsis phytochrome genes, PHYD and PHYE, are described and evidence is presented that, together with the previously described PHYA, PHYB and PHYC genes, the primary structures of the complete phytochrome family of this plant are now known. The PHYD- and PHYE-encoded proteins are of similar size to the other phytochrome apoproteins and show sequence similarity along their entire lengths. Hence, red/far-red light sensing in higher plants is mediated by a diverse but structurally conserved group of soluble photoreceptors. The proteins encoded by the PHYD and PHYE genes are more closely related to phytochrome B than to phytochromes A or C, indicating that the evolution of the PHY gene family in Arabidopsis includes an expansion of a PHYB-related subgroup. The PHYB and PHYD phytochromes show greater than 80% amino acid sequence identity but the phenotypes of phyB null mutants demonstrate that these receptor forms are not functionally redundant. The five PHY mRNAs are, in general, expressed constitutively under varying light conditions, in different plant organs, and over the life cycle of the plant. These observations provide the first description of the structure and expression of a complete phytochrome family in a higher plant.

Transgenic complementation of the hy3 phytochrome B mutation and response to PHYB gene copy number in Arabidopsis.

A recombinant PHYB minigene (mPHYB) consisting of the complete Arabidopsis PHYB cDNA sequence fused to 2.3 kb of upstream PHYB promoter sequence has been introduced into wild-type Arabidopsis and into a strain containing the Bo64 allele of the hy3 mutation. The Bo64 mutant has previously been shown to contain a nonsense mutation in the PHYB coding sequence. Transformation of this strain with the mPHYB gene results in complementation of all of the mutant phenotypic characteristics tested including hypocotyl length and hypocotyl cell size, response to end-of-day far-red light, leaf morphology, chlorophyll level, and flowering time. Presence of the mPHYB transgene in a wild-type genetic background causes exaggeration of this same set of phenotypic characteristics, indicating that these diverse photo-morphogenic responses are sensitive to the copy number of the PHYB gene. The transgene inserts in the Bo64(mPHYB) and WT(mPHYB) lines are shown to be single locus and single copy and the immunologically detectable level of phytochrome B is shown to vary linearly with PHYB gene copy number. These results demonstrate a complex role for phytochrome B in Arabidopsis photo-morphogenesis and suggest that the expression level of this phytochrome gene is an important determinant of the intensity of light-induced plant responses.

Labyrinthine functional tests and a case of midline cerebellar teratoma.

Electronystagmographic (ENG) examination may indicate clinically abnormal but nonspecific or even diagnostically deceptive results. Insights into the processes producing such observations sometimes can be suggested from correlations of findings to pathologic conditions in cases of demonstrated vestibular lesions. Such an opportunity occurred when unusual ENG results were obtained from a patient who had a history of equilibrium disturbances. The ENG revealed a direction-changing nystagmus in lateral head and body positions but not with the head similarly turned. Ocular counterroll measurements indicated hypofunctioning otolithic reactions despite normal vestibuloocular responses to calorizations. Radiologic follow-up showed a midline cerebellar tumor, later surgically confirmed, and found to be a rare teratoma. After partial excision and decompression, repeat tests revealed changes in both the positional nystagmus and ocular counterroll. These findings have implications for diagnosis, for mechanisms underlying ataxia, for inclusion of ocular counterroll testing in diagnostic evaluations, and for understanding the mechanisms underlying direction-changing positional nystagmus.

Ear-eye reflexes while riding in a car.

Persons with diminished or absent vestibulo-ocular reflexes (VORs) are thought to have difficulty with ocular stabilization during sufficiently rapid head movements. This suggests the hypothesis that labyrinthine defective individuals might have impaired visual abilities when in a moving motor vehicle. Ten licensed drivers likely to have abnormal VORs were recruited to assess this possibility. Their labyrinthine function was indexed by measurements of ocular counterroll responses to static head tilt and the nystagmus induced with caloric stimulations. Each person was evaluated on a visual task, i.e., answers to questions about alphanumeric information read from a stationary sign while sitting still and while riding in a car. Comparisons of vision scores tend to be lower in the moving than in the still condition. Furthermore, these decrements appear more closely related to the magnitudes of ocular counterroll than to the caloric reflexes. These results have implications concerning otolithic vs. semicircular canal function, vehicle operations by vestibular-impaired individuals, and evaluations of clinical treatments which employ vestibulotoxic drugs.

Detection of interaural phase shift between a subaudible and an audible tone.

Can a shift in interaural phase between a subthreshold signal and an audible contralateral probe tone affect perception of the probe? To obtain an answer, an 800-Hz tone was presented to both ears. The tone was presented continuously to one ear (-25 to + 10 dB SL) and in a sequence of four bursts per trial to the other ear (+ 10 dB SL). Interaural phase was reversed for either the second or the fourth burst in a 2 AFC task. Interaural phase-shift detection threshold (65% correct) varied with the intensity of the continuous signal; across subjects, this threshold varied from -21 to + 1 dB SL. When a 300-or 500-Hz masking tone was added to the ear with the continuous signal, phase-shift detection accuracy depended primarily upon the sensation level of the signal rather than its sound pressure level. These findings demonstrate temporal encoding at signal levels well below hearing threshold.

Interaural octave phase-shift detection and aural harmonic distortion.

A continuous 400-Hz tone (60-75 dB SPL) and 250-msec bursts of an 800-Hz tone (10 dB SL) were delivered dichotically. Four out of nine listeners were able to detect a 180 degree interaural phase shift. When a subaudible continuous 800-Hz tone was added to the ear with the 400-Hz tone, interaural phase-shift detection depended on the phase relation of the added tone to the 400-Hz tone. These results are shown to be consistent with the hypothesis of aural harmonic distortion.

Waardenburg's syndrome: a case report with CT scanning and cochleovestibular evaluation.

The Waardenburg's syndrome, a congenital ectodermal germ layer defect of autosomal dominant inheritance with variable phenotypic expressivity consists of 6 major characteristics: dystopia canthorum, broad nasal root, hypoplasia of the medial eyebrow, heterochromia irides, white forelock, and congenital deafness. Twelve additional characteristics have been reported including meningocele, atresia of the esophagus, and Hirshsprung's disease supporting an early hypothesis that the neural crest is the embryonic site linking defects of the cervical sympathic system with pigmentary abnormalities and inner ear anomalies. Congenital deafness reported in 20% of Waardenburg patients, and the most disabling characteristic, has been associated with ENG abnormalities in 30% of these deaf patients. A congenitally deaf young person with Waardenburg's syndrome is examined for basic cause of episodic vertiginous symptoms. Diagnostic assistance from previous investigations is lacking because they have not involved state-of-the-art functional evaluations or correlations of behavioral and radiographic findings. In this patient, high-resolution computerized tomography failed to demonstrate bony labyrinthine anomalies. Functional cochleovestibular analysis revealed a bilateral profound sensorineural hearing loss and a unilateral weakness in oculomotor responses to caloric stimulations. Repeated measurements of ocular counterroll show variations compatible with intermittent otolithic dysfunctioning. These findings are consistent with a diagnosis of an acquired active unilateral peripheral vestibular disorder. Some of the technical issues underlying the derivation of this conclusion are discussed.

Afterimage versus photographic ocular torsionometry.

Rotation of the eyeballs about the visual axis due to head tilt, counterocular torsion, can provide information relevant to the neuro-otologic status. Measurement of these reflexive movements, however, involves several difficulties. Two methods are examined: determinations by a behavioral afterimage technique are compared with those of a photographic method in the same subjects using a test-retest format. Results indicate that both procedures yield reliable measurements, and any differences tend to be inconsistent. Thus, the faster and less expensive afterimage technique appears suitable for many clinical purposes in assessing vestibulo-ocular function.

Aural harmonics: estimation consistency from tone-on-tone masking measurements.

Amplitudes of inaudible "subjective" signals are inferred from tone-on-tone masking measurements. Several methodological issues are involved, e.g., the problems of how many and which measurements to include. To explore such sampling questions, sets of masked thresholds using a 250-Hz masker and a 500-Hz maskee are determined at various phase angles. These measurements are distributed into subsamples of (n=) 13, 9, 7, 5, or 4 each, and repeated estimates of the second aural harmonic inferred. For n greater than 4, these estimates averaged to within +/- 1 dB of one another,, although individual estimates vary considerably with the smaller n's (to 13 dB with n=4). Sets of five consecutive measurements also were analyzed when the masker : maskee phase angle was incremented in 90 degrees steps through two complete periods (of the maskee). Again, the reestimates for individuals can vary by at least 6 dB, but the averages over seven subjects remain within 2 dB of each other. Apparently, for n's of six or more, it matters little which masker: maskee phase angles are used to estimate the aural harmonic amplitudes.

Loudness and the power series transformation. Preliminary model and measurements.

A nonlinear-algebraic approach to monaural intensity processing is proposed: the purpose is to integrate the conscious loudness attribute evoked by a pure tone with the power-series description of auditory distortion. The preliminary model consists of six postulates leading to a mathematical definition for loudness. To evaluate this equation, two task-specific sets of loudness judgments from each of seven subjects are examined. When linked to loudness-interval responses via the equisection assumption, the equation describes the behaviors quite well. Extrapolations into other intensity ranges predict similar responses with relatively slight overestimates. By relaxing the assumption that subjects will adjust the loudness proportions exactly as instructed, their ratio productions also can be described and predicted with sometimes surprising accuracy. Particularly striking is the prediction of overall levels and the curvilinearities of "doublings" from the "halvings". In addition, the theory proposes absolute loudness measurement, an explanation for the growth of loudness including the principle underlying Steven's Power Law, and might prove useful in examining some exceptions to this relationship. Several aspects of this model differ from traditional approaches to intensity processing, but it appears to warrent further critical evaluations.