Serum Glutamate Levels Correlate with Gleason Score and Glutamate Blockade Decreases Proliferation, Migration, and Invasion and Induces Apoptosis in Prostate Cancer Cells

During glutaminolysis, glutamine is catabolized to glutamate and incorporated into citric acid cycle and lipogenesis. Serum glutamate levels were measured in patients with primary prostate cancer (PCa) or metastatic castrate-resistant PCa (mCRPCa) to establish clinical relevance. The effect of glutamate-deprivation or blockade by metabotropic glutamate receptor 1 (GRM1)-antagonists was investigated on PCa cells’ growth, migration, and invasion to establish biological relevance

Identification of novel GRM1 mutations and single nucleotide polymorphisms in prostate cancer cell lines and tissues.

Metabotropic glutamate receptor 1 (GRM1) signaling has been implicated in benign and malignant disorders including prostate cancer (PCa). To further explore the role of genetic alterations of GRM1 in PCa, we screened the entire human GRM1 gene including coding sequence, exon-intron junctions, and flanking untranslated regions (UTRs) for the presence of mutations and single nucleotide polymorphisms (SNPs) in several PCa cell lines and matched tumor-normal tissues from Caucasian Americans (CAs) and African Americans (AAs). We used bidirectional sequencing, allele-specific PCR, and bioinformatics to identify the genetic changes in GRM1 and to predict their functional role. A novel missense mutation identified at C1744T (582 Pro > Ser) position of GRM1 gene in a primary AA-PCa cell line (E006AA) was predicted to affect the protein stability and functions. Another novel mutation identified at exon-intron junction of exon-8 in C4-2B cell line resulted in alteration of the GRM1 splicing donor site. In addition, we found missense SNP at T2977C (993 Ser > Pro) position and multiple non-coding mutations and SNPs in 3'-UTR of GRM1 gene in PCa cell lines and tissues. These novel mutations may contribute to the disease by alterations in GRM1 gene splicing, receptor activation, and post-receptor downstream signaling

Identification of Novel <i>GRM1</i> Mutations and Single Nucleotide Polymorphisms in Prostate Cancer Cell Lines and Tissues

<div><p>Metabotropic glutamate receptor 1 (GRM1) signaling has been implicated in benign and malignant disorders including prostate cancer (PCa). To further explore the role of genetic alterations of <i>GRM1</i> in PCa, we screened the entire human <i>GRM1</i> gene including coding sequence, exon-intron junctions, and flanking untranslated regions (UTRs) for the presence of mutations and single nucleotide polymorphisms (SNPs) in several PCa cell lines and matched tumor-normal tissues from Caucasian Americans (CAs) and African Americans (AAs). We used bidirectional sequencing, allele-specific PCR, and bioinformatics to identify the genetic changes in <i>GRM1</i> and to predict their functional role. A novel missense mutation identified at C1744T (582 Pro > Ser) position of <i>GRM1</i> gene in a primary AA-PCa cell line (E006AA) was predicted to affect the protein stability and functions. Another novel mutation identified at exon-intron junction of exon-8 in C4-2B cell line resulted in alteration of the <i>GRM1</i> splicing donor site. In addition, we found missense SNP at T2977C (993 Ser > Pro) position and multiple non-coding mutations and SNPs in 3′-UTR of <i>GRM1</i> gene in PCa cell lines and tissues. These novel mutations may contribute to the disease by alterations in <i>GRM1</i> gene splicing, receptor activation, and post-receptor downstream signaling.</p></div

Genotype status of mutations and SNPs identified in <i>GRM1</i> gene in matched prostate tumor-normal tissues.

a<p>DSSC: Downstream to stop codon.</p

Genotype status of non-synonymous and synonymous mutations and polymorphisms identified in <i>GRM1</i> gene in prostate cancer cell lines.

a<p>Novel mutations identified in our study.</p>b<p>Mis: Missense mutation.</p

Schematic presentation of the identified mutations and polymorphisms of full-length <i>GRM1</i> gene in prostate cancer cell lines and tissues.

<p>Schematic presentation of the identified mutations and polymorphisms of full-length <i>GRM1</i> gene in prostate cancer cell lines and tissues.</p

Allele-specific PCR genotyping of novel missense mutation identified in exon-8 of <i>GRM1</i> gene in prostate tumors.

<p>Presence of C-allele showed amplification of a 439 bp fragment and presence of T-allele showed amplification of a 267 bp fragment. A nonspecific fragment of 706 bp was amplified in all samples.</p

Bioinformatics prediction of splicing site motif (donor/acceptor).

<p>Twenty one base pair sequence flanking of mutation site with wild type (<b>A</b>) and mutant allele (<b>B</b>) was tested for prediction of splicing site motif (donor or acceptor).</p

Genotype status of downstream non-coding polymorphisms identified in <i>GRM1</i> gene in prostate cancer cell lines.

a<p>DSSC: Downstream to stop codon.</p

Location of the novel identified mutations and polymorphisms in different domains of GRM1 receptor.

<p>Location of the novel identified mutations and polymorphisms in different domains of GRM1 receptor.</p