Analysis of the Golden Syrian Hamster Anterior Pituitary Gland Proteome by Ion Trap Mass Spectrometry

Abstract

We utilized mass spectrometry (MS) and bioinformatics to investigate the proteome of the anterior pituitary gland (AP). Subcellular fractions of APs from 2-month-old male Golden Syrian hamsters were prepared for protein denaturation, treatment with trypsin and analyses utilizing micro liquid chromatography MS/MS and the database search software SEQUEST. In the nuclear, non-nuclear 100,000 × g and cytosolic fractions we identified 76, 52 and 52 different proteins, respectively. A total of 145 distinct proteins were detected. We identified growth hormone, prolactin, pro-opiomelanocortin, the α-subunit for the glycoprotein hormones, luteinizing hormone-β and follicle-stimulating hormone-β. Groups of other identified proteins included hormone processing, secretion granule associated, non-hormonal endoplasmic reticulum associated, calcium binding, protein kinase C associated histone and non-histone chromosomal material, other RNA-binding, splicing factors, heterogeneous nuclear ribonucleoproteins, helicases, lamins, microfilament associated, microtubule associated, adenosine triphosphate and guanosine diphosphate associated, keratins, lysosomal, ribosomal, enzymes in glycolysis and the tricarboxylic and pentose phosphate paths, glutathione associated, transmethylation, catabolic and unknown protein products as well as blood hemoglobins. Proteins previously not reported in the AP, such as fertility protein SP22, were identified. The proteins identified in the present study form a foundation for defining the proteome in normal adult male AP.

Mouse Anterior Pituitary Gland: Analysis by Ion Trap Mass Spectrometry

“…), the chest cavity was opened and the right atrium cut, and the animals were perfused through the left ventricle with PBS perfusion solution (2.7 m M Na 2 HPO 4 , 1.5 m M KH 2 PO 4 , 154 m M NaCl; pH 7.2) for approximately 2 min. After removing the cranial vault and brain to visualize the pituitary gland, the posterior lobe (pars intermedia and pars nervosa) of the pituitary gland was separated from the AP in situ under magnification and with the use of fine forceps as described for hamster [1]. The AP (approx.…”

“…Samples were digested with trypsin prior to rather than after reduction-alkylation to avoid an additional step of dialysis usually required between reduction-alkylation and tryptic digestion. This protocol was used successfully in previous studies [1, 2]. …”

“…We recently utilized micro liquid chromatography tandem mass spectrometry (LC/MS/MS) and the database search software SEQUEST to identify a total of 145 different proteins in subcellular fractions of the anterior pituitary gland (AP) of the Golden Syrian hamster [1]. The proteins included hamster growth hormone (GH), prolactin (PRL), pro-opiomelanocortin (POMC) the precursor of adrenocorticotropic hormone (ACTH), glycoprotein hormones α-subunit, and luteinizing hormone (LH)-β as well as rat follicle-stimulating hormone (FSH)-β. Thyroid-stimulating hormone (TSH)-β was not identified.…”

“…In the present study we utilized methodology similar to that employed to analyze the hamster AP [1] to investigate the AP proteome in the mouse. We identified 115 different proteins in the mouse AP.…”

“…We identified 115 different proteins in the mouse AP. Approximately half of these proteins had not been identified in the AP of the hamster [1], including hormone-processing and secretion granule-associated proteins. Between the two species, over 200 different proteins have been identified in the AP of young adult males, providing some interesting observations and comparisons between the two species.…”