Evolutionary Analysis of the Steroidogenic Acute Regulatory Protein-Related Lipid Transfer Domain and Its Response to Salt Stress In Vitis Vinifera

This study aimed to enhance the understanding of the steroidogenic acute regulatory protein-related lipid transfer (START) domain in Vitis vinifera. A total of 23 members of the VvSTARD gene family were found, which could be divided into five groups. The analyses of the gene codon preference, selective pressure, and tandem replication events of the VvSTARD, AtSTARD, and OsSTARD genomes indicated that tandem replication events occured in grapes, Arabidopsis, and rice genomes. Eight lipid transporter proteins were found in the tertiary structure of the STARD gene family in grapes. The analysis of the expression profiles of the three species microarrays showed that the expression sites of the STARD gene and the response to abiotic stress in the same subgroup had similar characteristics. In addition, quantitative real-time polymerase chain reaction (qRT-PCR) was used to analyze the expression of the STARD gene family in grape leaves in response to different hormones and abiotic stresses, and the obtained results were the same as those predicted by the cis-elements and the expression profiles. Furthermore, 35S:STARD5:EGFP was successfully constructed to verify the subcellular prediction results, and the results showed that STARD5 was located in the nucleus. Through the identification of salt tolerance of transgenic tomato, STARD5 was found to regulate the salt stress of plants. Collectively, these data indicated that the VvSTARD gene family plays an important role in response to salt stress.

Incomplete Pattern of Steroidogenic Protein Expression in Functioning Adrenocortical Carcinomas

Autonomous steroid secretion is a common feature of adrenocortical carcinomas (ACC), although not always clinically evident owing to inefficient steroidogenesis with increased release of steroid precursors. Our study aim was to analyze the expression profile of four key proteins involved in the steroidogenesis cascade, in different adrenocortical tumors. Expression of proteins involved in steroidogenesis, namely steroidogenic acute regulatory protein (StAR), 11β-hydroxylase (CYP11B1), aldosterone synthase (CYP11B2) and 17α-hydroxylase (CYP17A1), were analyzed by immunohistochemistry in ACC (n = 14), adenomas presenting with Cushing’s syndrome (ACAc) (n = 11) and clinically non-functioning adenomas (ACAn) (n = 15). A percentage of the stained area for each protein was analyzed using ImageJ software for computerized morphometric quantification. CYP11B1, StAR and CYP17A1 expression were significantly lower in ACC when compared to ACAc. In addition, ACC presented co-staining cells for CYP11B1 and CYP11B2. CYP11B1 was the steroidogenic enzyme with the most discriminative power to distinguish ACC from ACAc, with a sensitivity of 100%, specificity of 92%, and an expression higher than 4.44%, indicating the presence of a cortisol secreting adenoma. ACC depicts an incomplete pattern of steroidogenic protein expression, with decreased CYP11B1 and CYP17A1, which could explain the predominant secretion of steroid precursors.

Molecular basis for the recognition of steroidogenic acute regulatory protein by the 14-3-3 protein family

ABSTRACTSteroidogenesis in adrenals and gonads starts from cholesterol transport to mitochondria by the steroidogenic acute regulatory protein STARD1, containing a mitochondrial import sequence followed by a cholesterol-binding START domain. Although mutations in this protein have been linked to lipoid congenital adrenal hyperplasia, the mechanism of steroidogenesis regulation by the STARD1 remains debatable, hypothetically involving a molten-globule structural transition and interaction with 14-3-3 proteins. We show that, while the isolated START domain does not interact with 14-3-3, interaction is enabled by STARD1 phosphorylation at Ser57, close to the mitochondrial peptide cleavage site. Biochemical analysis of the STARD1 affinity towards 14-3-3 and crystal structures of 14-3-3 complexes with Ser57 and Ser195 phosphopeptides, suggest distinct roles of site-specific phosphorylations in recruiting 14-3-3, to modulate STARD1 activity, processing and import to mitochondria. Phosphorylation at Ser195 creates a unique conditional site, that could only bind to 14-3-3 upon partial unfolding of the START domain.