Antizyme inhibitor 1 (Azin1), a positive regulator of cellular polyamines, is induced by various proliferative stimuli and repressed by polyamines. It has been reported that the translational repression of Azin1 by polyamines involves an upstream open reading frame on the mRNA, but little has been known about polyamine effect on its transcription or splicing. We found multiple forms of Azin1 transcripts formed by alternative splicing and initiation of transcription from putative alternative start sites. One of the novel splice variants, Azin1-X, has a premature termination codon on 5′ extension of exon 7, encodes a C-terminal truncated form of protein (Azin1ΔC), and is subject to nonsense-mediated mRNA decay. 2-Difluoromethylornithine (DFMO), an inhibitor of polyamine synthesis, increased both transcription from the canonical transcription start site and the ratio of the full-length mRNA to Azin1-X mRNA, whereas polyamines show the opposite effect. Thus, polyamines regulate two novel steps of Azin1 expression, namely the transcription and a particular splicing pattern, both of which may affect the level of mRNA encoding the full-length active Azin1 protein.