Two potent nuclear localization signals in the gut-enriched Krüppel-like factor define a subfamily of closely related Krüppel proteins.

Janiel M Shields,Vincent W Yang

doi:10.1074/jbc.272.29.18504

Abstract

The gut-enriched Krüppel-like factor (GKLF) is a newly identified transcription factor that contains three C2H2 Krüppel-type zinc fingers. Previous immunocytochemical studies indicate that GKLF is exclusively localized to the nucleus. To identify the nuclear localization signal (NLS) within GKLF, cDNA constructs with various deletions in the coding region of GKLF were generated and analyzed by indirect immunofluorescence in transfected COS-1 cells. In addition, constructs fusing regions representing putative NLSs of GKLF to green fluorescent protein (GFP) were generated and examined by fluorescence microscopy in similarly transfected cells. The results indicate that GKLF contains two potent, independent NLSs: one within the zinc fingers and the other in a cluster of basic amino acids (called 5' basic region) immediately preceding the first zinc finger. In comparison, putative NLSs within the zinc fingers and the 5' basic region of a related Krüppel protein, zif268/Egr-1, are relatively less efficient in their ability to translocate GFP into the nucleus. A search in the protein sequence data base revealed that despite the existence of numerous Krüppel proteins, only two, the lung Krüppel-like factor (LKLF) and the erythroid Krüppel-like factor (EKLF), exhibit similar NLSs to those of GKLF. These findings indicate that GKLF, LKLF, and EKLF are members of a subfamily of closely related Krüppel proteins.

Highlights

Various mechanisms that are responsible for nuclear localization of eukaryotic transcription factors have been proposed
These results indicate that the 5Ј basic region of gut-enriched Kruppel-like factor (GKLF) is both necessary and sufficient for nuclear localization
Significant progress has been made toward the understanding of the mechanisms that mediate localization of proteins to the nucleus. This is in part due to the availability of data bases containing amino acid sequences of a large number of transcription factors

Summary

Introduction

Various mechanisms that are responsible for nuclear localization of eukaryotic transcription factors have been proposed. Many putative NLSs are present in close proximity to the DNA-binding domains of transcription factors, exemplified by the bZIP proteins c-Fos and c-Jun, and the bHLH proteins Myc, Max, and Myo D1 [3] This conserved arrangement seems to suggest that DNA-binding motifs and nuclear localization signals may have coevolved. The primary amino acid sequence in the zinc finger region of GKLF exhibits a high degree of identity with several previously identified Kruppel proteins, including lung Kruppellike factor (LKLF [7]), erythroid Kruppel-like factor (EKLF [8]), and basic transcription element-binding protein 2 (BTEB2 [9]). Our results suggest that the Kruppel family of transcription factors can further be divided into subfamilies based on the sequences required for nuclear localization

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