Allopolyploidy is a peculiar process entailing the cohabitation of two (or more) divergent genomes. Consequences on plant genomes are varied and can utlimately alter gene expression and regulatory interactions. Most studies have explored polyploid expression evolution in experimental controlled conditions. Here, we analyzed global gene expression variation in natural populations of the Spartina polyploid complex including hexaploid parental species (S. maritima and S. alterniflora), two F1 hybrids (S. × townsendii and S. × neyrautii) and the allododecaploid S. anglica. In situ sampling and quantitative PCR were performed for comparing global expression of candidate genes involved in responses to abiotic stresses, lignin and cellulose metabolisms between five Spartina taxa. Illumina sequencing datasets and dedicated bioinformatic pipelines were employed to explore sequence heterogeneity in these highly duplicated genomes. Levels of gene expression were significantly higher in S. alterniflora (compared to the other hexaploid parent S. maritima) for seven of the analyzed genes. Effects of both hybridization and polyploidization are detected and consistent with previous global transcriptome analyses performed on Spartina plants grown in controlled conditions. Duplicated copies present in the hybrids and the allododecaploid were successfully assigned to either one of the parental genomes. Phylogenetic analyses identified for each of the parental hexaploid species, the presence of two distinct clades including two or more expressed copies. We provide here a comprehensive gene expression study based on individuals sampled in their natural habitat and detected the superimposed effect of environmental heterogeneity, hybridization and allopolyploidy.