Despite advances in the understanding of the pathogenesis of salivary gland neoplasms (SGN), the molecular pathways associated with enhanced tumor growth and cell survival remain to be established. The aim of the present study was to investigate whether
TP53
mutations are relevant to SGN pathogenesis and if they impact on p53 protein expression. The study included 18 benign and 18 malignant SGN samples. Two polymorphic microsatellite markers at the
TP53
genetic locus were chosen to assess loss of heterozygosity (LOH) in the samples that had matched normal DNA. The
TP53
exons 2–11 were amplified by PCR, and all of the products were sequenced. Reverse transcription-PCR of the
TP53
open reading frame (ORF) was carried out in the samples that had fresh tissue available, and immunohistochemistry for the p53 protein was performed in all samples.
TP53
LOH was only found in two pleomorphic adenomas. We found two missense mutations in exon 7 (one in a pleomorphic adenoma and the other in a polymorphous low grade adenocarcinoma), another in exon 8 (in a carcinoma ex pleomorphic adenoma) and a fourth missense mutation in exon 10 (in a mucoepidermoid carcinoma). In addition, a nonsense mutation was found in exon 8 of an adenoid cystic carcinoma. Several intronic and exonic SNPs were detected. Although almost all of the malignant samples were immunopositive for p53, approximately 37% of the benign samples were positive, including the sample harboring the missense mutation and one of the samples that showed LOH. The complete
TP53
ORF could be amplified in all samples analyzed, including the IHC negative samples, the samples showing LOH and one sample displaying a missense mutation. In summary, our results show that
TP53
mutations are not a frequent event in SGN and that p53 immunopositivity might not be associated with sequence mutations in SGN.
In trypanosomatids, regulation of
gene expression occurs mainly
at the posttranscriptional level, and RNA-binding proteins (RBPs)
are key players in determining the fates of transcripts. RBPs are
targets of protein arginine methyltransferases (PRMTs), which posttranslationally
regulate the RNA-binding capacity and other RBP interactions by transferring
methyl groups to arginine residues (R-methylation). Herein, we functionally
characterized the five predicted PRMTs in Leishmania braziliensis by gene knockout and endogenous protein HA tagging using CRISPR/Cas9
gene editing. We report that R-methylation profiles vary among Leishmania species and across L. braziliensis lifecycle stages, with the peak PRMT expression occurring in promastigotes.
A list of PRMT-interacting proteins was obtained in a single coimmunoprecipitation
assay using HA-tagged PRMTs, suggesting a network of putative targets
of PRMTs and cooperation between the R-methylation writers. Knockout
of each L. braziliensis PRMT led to significant
changes in global arginine methylation patterns without affecting
cell viability. Deletion of either PRMT1 or PRMT3 disrupted most type
I PRMT activity, resulting in a global increase in monomethyl arginine
levels. Finally, we demonstrate that L. braziliensis PRMT1 and PRMT5 are required for efficient macrophage infection
in vitro, and for axenic amastigote proliferation. The results indicate
that R-methylation is modulated across lifecycle stages in L. braziliensis and show possible functional overlap
and cooperation among the different PRMTs in targeting proteins. Overall,
our data suggest important regulatory roles of these proteins throughout
the L. braziliensis life cycle, showing that
arginine methylation is important for parasite–host cell interactions.
Malignant salivary neoplasms showed a higher frequency of the allele encoding Arg and a higher frequency of the Arg/Arg genotype. However, the different genotypes did not impact the transcription of genes involved in apoptosis.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.