CD44 pre-mRNA includes 20 exons, among which exons1-5 (C1-C5) and exons16-20 (C6-C10) are constant exons, whereas exons 6-15 (V1-V10) are variant exons. V6 exon containing isoforms has been known to be implicated in tumor cell invasion and metastasis. In the present study, we performed SR protein screen for CD44 V6 splicing using overexpression and lentivirus-mediated shRNA treatment. Using CD44 V6 minigene, we demonstrate that increased SRSF3 and SRSF4 expression do not affect V6 splicing, but increased expression of SRSF1, SRSF6 and SRSF9 inhibit V6 splicing significantly. In addition, using constitutive exon specific primer set, we could not detect alteration of CD44 splicing after SR protein-targeting shRNA treatment. However, using V6 specific primer, we identified that reduced SRSF2 expression significantly reduced V6 isoform, but increased V6-10 and V6,7-10 isoforms. Our results indicate that SR proteins are important regulatory proteins for CD44 V6 splicing.