Lysosomal membranes are enriched in extensively glycosylated transmembrane proteins, LAMP-1 and LAMP-2. LAMP-1 proteins have been characterized from several mammalian species and from chickens, but no non-mammalian homologues of LAMP-2 have been described, and no splice variants of either protein have been reported. Here we report the characterization of three cDNA clones encoding chicken LAMP-2. The nucleotide sequences of the cDNAs diverge at their 3' ends within the open reading frame, resulting in sequences that code for three different transmembrane and cytoplasmic domains. Southern analysis suggests that a single gene encodes the common region of chicken LAMP-2. The position of the divergence and the identity of the common sequence are consistent with alternative splicing of 3' exons. Analysis of the mRNAs present in adult chicken tissues suggests tissue-specific expression of the three chicken LAMP-2 variants, with LAMP-2b expressed primarily in the brain. The cytoplasmic domain of LAMP-type proteins contains the targeting signal for directing these molecules to the lysosome. Using chimeras consisting of the lumenal domain of chicken LEP100 (a LAMP-1) and the transmembrane and cytoplasmic domains of the LAMP-2 variants, we demonstrate in transfected mouse L cells that all three LAMP-2 carboxyl-terminal regions are capable of targeting the chimeric proteins to lysosomes. Levels of expression, subcellular distribution, and glycosylation of the LAMP proteins have all been shown to change with differentiation in mammalian cells and to be correlated with metastatic potential in certain tumor cell lines. Alternative splicing of the LAMP-2 transcript may play a role in these changes.