José A. Díaz

Exome sequencing identifies MAX mutations as a cause of hereditary pheochromocytoma

Hereditary pheochromocytoma (PCC) is often caused by germline mutations in one of nine susceptibility genes described to date, but there are familial cases without mutations in these known genes. We sequenced the exomes of three unrelated individuals with hereditary PCC (cases) and identified mutations in MAX, the MYC associated factor X gene. Absence of MAX protein in the tumors and loss of heterozygosity caused by uniparental disomy supported the involvement of MAX alterations in the disease. A follow-up study of a selected series of 59 cases with PCC identified five additional MAX mutations and suggested an association with malignant outcome and preferential paternal transmission of MAX mutations. The involvement of the MYC-MAX-MXD1 network in the development and progression of neural crest cell tumors is further supported by the lack of functional MAX in rat PCC (PC12) cells and by the amplification of MYCN in neuroblastoma and suggests that loss of MAX function is correlated with metastatic potential.

Genetics of pheochromocytoma and paraganglioma in Spanish patients.

CONTEXT The presence of familial history in pheochromocytoma/paraganglioma patients, including syndromic antecedents, leads in the majority of cases to a positive genetic testing for mutations in one of the major susceptibility genes described so far. Furthermore, it has been reported that in the absence of familial antecedents, about 11-24% of patients also carry a mutation in one of these related genes. In these cases, other clinical aspects like bilaterality, multiplicity, location of the tumors, or age at onset can help to recognize the underlying genes involved. OBJECTIVE The objective of the study was to discuss clinical criteria helpful in the genetic diagnosis, placing special emphasis on apparently sporadic cases. DESIGN Two hundred thirty-seven nonrelated probands were analyzed for the major susceptibility genes: VHL, RET, SDHB, SDHC, and SDHD. Genetic characterization included both point mutation analysis and gross deletions in the SDH genes performed by multiplex PCR. RESULTS As expected, all syndromic probands were genetically diagnosed with a mutation affecting either RET or VHL. A total of 79.1% (19 of 24) and 18.4% (31 of 168) of patients presenting with either nonsyndromic familial antecedents or apparently sporadic presentation were found to carry a mutation in one of the susceptibility genes. Finally, we found a Spanish founder effect for two mutations: SDHB c.166_170delCCTCA and SDHD c.129G>A. CONCLUSIONS Germline mutations are rare in apparently sporadic probands diagnosed after age 40 yr (3.9% in our series) and mainly involve SDHB. Therefore, we recommend prioritizing SDHB genetic testing in patients developing isolated tumors at any age, especially those with extraadrenal location or malignant behavior.

Covariation of Thermal Biology and Foraging Mode in Two Mediterranean Lacertid Lizards

Body temperatures, heat exchange rates, behavioral thermoregulation, and movement behavior (as an index of foraging mode) were studied in two widely distributed, medium-sized lacertid lizards (Acanthodactylus erythrurus and Psammodromus algirus). P. algirus mainly inhabits broad-leaved forests, whereas A. erythrurus prefers open, sandy areas with sparsely distributed vegetation. These habitat preferences parallel differences between the areas in which both genera presumably originated: Eurosaharian xeric steppes with high operative temperatures (T e ) for Acanthodactylus, and Mediterranean open forests with lower T e for Psammodromus. Field observations showed that percentage of time spent basking and basking rate (number of basks per minute) were negatively related to T e , although average bask duration was not. Percentage of time spent moving, moving rate (number of moves per minute), and the average duration of individual moves were inversely related to T e and were higher in P. algirus. The percentage of total locomotion time that was spent moving in the shade was also higher in P. algirus. Behavioral thermoregulation strategies differed in a laboratory thermogradient, where P. algirus basked more often and for shorter periods and selected warmer patches than did A. erythrurus. Selected body temperatures (T b ) in a laboratory thermogradient were significantly higher in A. erythrurus than in P. algirus. Shade Seeking T b was higher in A. erythrurus, but Resume Basking T b did not differ significantly between the two species. Heating and cooling rates also differed in the two species: A. erythrurus warmed more slowly and cooled faster than did P. algirus. Our data support the existence of a complex syndrome that combines aspects of the behavior, physiology, and ecology of both species. The thermal consequences of inhabiting a certain type of habitat can be counterbalanced by behavioral and physiological means that, in turn, affect movement and, hence, foraging behavior. Thus, the more active species (P. algirus) heated faster, cooled more slowly, and basked more often but for shorter periods and at warmer patches than the less active species (A. erythrurus).

Regional distribution of a Mediterranean lizard: influence of habitat cues and prey abundance

We studied the distribution of a common Medi- terranean lacertid lizard, Psammodromus algirus (L.) 1758, on nineteen sites within a regional gradient of homoge- neous yet contrasted habitats. This scale was large enough to allow line-transect estimates of lizard abundance, which were related to quantitative (and when possible multivari- ate) measurements of the structure and floristic composition of vegetation, the abundance of arthropod prey, the relative density of other lizard species, and the climatic data obtained from nearby meteorological stations. Neither the climate not the abundance of other lizards seemed to condi- tion the quantitative distribution of the species. The positive influence of broad-leaved forests on the abundance of P. more directly related to the ecology of lizards than floristic composition per se. Thus, population levels were most highly correlated with the cover of shrubs over 20 cm in height, and once this stnictural requirement was met, they increased with the abundance of potential prey (itself condi- tioned by vegetation cover at the ground leve1 and litter cover). We suggest that our results should be interpreted in the context of thermoregulatory, predator avoidance and movement minimization strategies whose influence on sur- vival, and hence abundance, could probably be applied to other insectivorous lizards from temperate zones.

Gross SDHB deletions in patients with paraganglioma detected by multiplex PCR: A possible hot spot?

Pheochromocytoma and paraganglioma are rare neuroendocrine tumors that arise in the adrenal medulla and the extra‐adrenal paraganglia, respectively. Inheritance of these tumors is mainly a result of mutations affecting the VHL, RET, NF1, and SDH genes. Germ‐line mutations of the SDH genes have been found to account for nearly 10% of apparently sporadic cases. Nevertheless, alterations other than point mutations have not yet been well characterized. In this study, we investigated the frequency of gross SDH deletions in 24 patients who tested negative for point mutations and had at least one of the recommended features for genetic testing. For this purpose, we used a technique that is easy to implement in the lab to specifically detect gross deletions affecting SDHB, SDHC, and SDHD. We identified 3 heterozygous SDHB deletions (3/24) in 3 independent cases with paraganglioma: 1 whole SDHB deletion and 2 deletions exclusively affecting exon 1. These latter mutations match the unique gross deletion previously reported, indicating this region could be a hot spot for gross SDHB deletions. It seems likely that these alterations can account for a considerable number of both familial and apparently sporadic paraganglioma cases. Although this is the first report describing the presence of gross deletions in patients with apparently sporadic paragangliomas, the extra‐adrenal location of the tumor seems to constitute a determining factor for whether to include these patients in genetic testing for gross deletions in the SDHB gene.

Recommendations for somatic and germline genetic testing of single pheochromocytoma and paraganglioma based on findings from a series of 329 patients

Background Nowadays, 65–80% of pheochromocytoma and paraganglioma (PPGL) cases are explained by germline or somatic mutations in one of 22 genes. Several genetic testing algorithms have been proposed, but they usually exclude sporadic-PPGLs (S-PPGLs) and none include somatic testing. We aimed to genetically characterise S-PPGL cases and propose an evidence-based algorithm for genetic testing, prioritising DNA source. Methods The study included 329 probands fitting three criteria: single PPGL, no syndromic and no PPGL family history. Germline DNA was tested for point mutations in RET and for both point mutation and gross deletions in VHL, the SDH genes, TMEM127, MAX and FH. 99 tumours from patients negative for germline screening were available and tested for RET, VHL, HRAS, EPAS1, MAX and SDHB. Results Germline mutations were found in 46 (14.0%) patients, being more prevalent in paragangliomas (PGLs) (28.7%) than in pheochromocytomas (PCCs) (4.5%) (p=6.62×10−10). Somatic mutations were found in 43% of those tested, being more prevalent in PCCs (48.5%) than in PGLs (32.3%) (p=0.13). A quarter of S-PPGLs had a somatic mutation, regardless of age at presentation. Head and neck PGLs (HN-PGLs) and thoracic-PGLs (T-PGLs) more commonly had germline mutations (p=2.0×10−4 and p=0.027, respectively). Five of the 29 metastatic cases harboured a somatic mutation, one in HRAS. Conclusions We recommend prioritising testing for germline mutations in patients with HN-PGLs and T-PGLs, and for somatic mutations in those with PCC. Biochemical secretion and SDHB-immunohistochemistry should guide genetic screening in abdominal-PGLs. Paediatric and metastatic cases should not be excluded from somatic screening.

Seasonal variation of gonadal development, sexual steroids, and lipid reserves in a population of the lizard Psammodromus algirus

We studied the seasonal variations of reproductive characteristics in a wild population of the lacertid lizard Psammodromus algirus by measuring gonadal weight, germinal epithelium height, sexual steroids (testosterone, estradiol, and progesterone), and lipid reserves (in fat bodies and liver) in three phases of the reproductive cycle: early breeding (late April), late breeding (mid-June), and post-breeding season (late July). In males, testis size, germinal epithelium, and testosterone plasma levels were high in the breeding season and decreased dramatically in July. However, spermatogenesis (maximum in June) was slightly delayed with respect to testosterone secretion (peak in April), suggesting that a previous increase in testosterone levels could be necessary to stimulate the initial stages of spermatogenesis

A Comparative Study of the Relation between Heating Rates and Ambient Temperatures in Lacertid Lizards

It is well known that lizard species from thermally diverse areas differ in their use of thermoregulatory behaviors as a means of making short-term adjustments to differences in ambient conditions. In contrast, the extent of long-term adjustments in thermal physiology is poorly documented and still under debate. In this study we report a clear interspecific relationship between heating rates, measured under standardized laboratory conditions, and environmental temperatures within a small clade of lacertid lizards. Phylogenetically based ANCOVAs demonstrate that species with a northern or montane distribution warm at a faster mass-specific rate than do species from a southern (Mediterranean) climatic zone. Correlational analyses, using phylogenetically independent contrasts, confirm that continuous among-species variation in mass-specific heating rates is negatively related to clinal differences in environmental temperatures. A reduction of the time spent warming, associated with higher heating rates, should be especially advantageous for lizards from cool climates, which bask for large amounts of time. Thus, the observed relation between heating rates and climatic conditions could be a result of adaptive evolution. The behaviorally selected preferred temperatures were lower, but not significantly so, in the northern or montane species set; interspecific correlations with ambient temperatures were positive but not significant. However, the divergence of heating rates and the statistically nonsignificant difference in preferred temperatures have similar impacts on variation in the estimated amount of time spent heating. Thus, differences in both heating rate, a physiological trait, and preferred temperature, a behavioral characteristic, contribute equally to variation in heating times.

TEMPORAL PATTERNS OF BASKING BEHAVIOUR IN A MEDITERRANEAN LACERTID LIZARD

1. I studied the thermoregulatory behaviour of the diurnal basker Psammodromus algirus (Reptilia, Lacertidae) in the mosaic of sunshade patches of a Mediterranean evergreen forest. 48 behavioural sequences associated with measurements of the thermal environment provided data on the percentages of time spent basking by focal animals, the duration of individual basking periods and the frequency of basking. 2. The percentage of time basking decreased linearly from the onset of the activity period until midday. The average duration of basking periods had a pronounced decrease after 0700 h and remained constant throughout the rest of the morning, whereas the basking rates, that were minimal before 0700 h, reached a peak between 0700 and 0800 h followed by a progressive fall that continued until noon. 3. In order to explain these behavioural patterns, I experimentally determined the effects of environmental variables (air and ground temperature and solar angle) on the heating and cooling rates of captive lizards. The resulting regression models were then used to predict the heat exchange rates of the observed free-ranging animals. 4. The moment at which heating rates should equal and then exceed cooling rates, appeared to set off the behavioural shift towards a larger frequency of shorter basking periods. Under these circumstances, the apparently random movement of a shuttling heliotherm with respect to sun and shade might still be of thermoregulatory significance.

How Much Variation in the Molt Duration of Passerines Can Be Explained By the Growth Rate of Tail Feathers

ABSTRACT. In birds, molt duration is an important trait that can affect plumage functionality and, consequently, the fitness of individuals. However, knowledge about the factors that affect variation in molt speed is sparse, mostly because of the methodological difficulties of studying avian molt. We used a ptilochronology-based approach to estimate the rate at which tail feathers were produced during molt to shed light on the relationship between molt duration and feather growth rate. For that purpose, we used three data sets. First, we tested whether the average molt durations of 22 passerine species were correlated with the mean growth rates of their feathers, using both conventional and phylogenetically corrected statistical procedures. Second, we explored this same association among captive Great Tits (Parus major). And third, we took advantage of the biannual complete molt of Willow Warblers (Phylloscopus trochilus) to examine whether the feathers synthesized during their short summer molt grew faster than those produced during their long winter molt. Feather growth rates were negatively correlated with molt duration in all analyses, revealing that molt duration can be estimated from the growth rate of a single feather. However, predictive power was limited by the fact that molt duration is modulated mainly by molt intensity, which seems to be correlated with ecological constraints in our interspecific approach. We also discuss the implications of our results for the evolution of molt duration, and the potential application of ptilochronology in its study.